                         SEQUENCE LISTING

<110>  CENTRE NATIONAL DE LA RECHERCHE SCIENTIFIQUE (CNRS)
 
<120>  GENETICALLY MUTATED AEQUORIN-BASED BIOSENSORS WITH EXTRA 
       SENSITIVITY TO CALCIUM AND IMPROVED BIOLUMINESCENCE INTENSITY

<130>  B370936D34643

<150>  EP 15306738.4
<151>  2015-10-30

<160>  25    

<170>  PatentIn version 3.5

<210>  1
<211>  189
<212>  PRT
<213>  Aequorea aequorea


<220>
<221>  SITE
<222>  (1)..(189)
<223>  wild-type aequorin

<400>  1

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Gln Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  2
<211>  12
<212>  PRT
<213>  Aequorea aequorea


<220>
<221>  SITE
<222>  (1)..(12)
<223>  aequorin EF Hand

<400>  2

Asp Val Asn His Asn Gly Lys Ile Ser Leu Asp Glu 
1               5                   10          


<210>  3
<211>  12
<212>  PRT
<213>  Aequorea aequorea


<220>
<221>  SITE
<222>  (1)..(12)
<223>  aequorin EF Hand

<400>  3

Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
1               5                   10          


<210>  4
<211>  12
<212>  PRT
<213>  Aequorea aequorea


<220>
<221>  SITE
<222>  (1)..(12)
<223>  aequorin EF Hand

<400>  4

Asp Ile Asp Glu Ser Gly Gln Leu Asp Val Asp Glu 
1               5                   10          


<210>  5
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  5

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Asp Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  6
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  6

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Thr Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  7
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  7

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Gly Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  8
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  8

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Glu Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  9
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  9

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asp Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Gln Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  10
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  10

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Asp Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Gln Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  11
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  11

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Gln Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Thr Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  12
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  12

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Asp Gly Gln Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  13
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  13

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asp Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Asp Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  14
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  14

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asp Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Thr Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  15
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  15

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Asp Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Asp Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  16
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  16

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Asp Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Thr Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  17
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  17

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Asp Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Thr Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  18
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  18

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Thr Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Thr Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  19
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  19

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Asp Gly Asp Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  20
<211>  189
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  20

Ser Lys Leu Thr Ser Asp Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
1               5                   10                  15      


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
            20                  25                  30          


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
        35                  40                  45              


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
    50                  55                  60                  


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
65                  70                  75                  80  


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
                85                  90                  95      


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
            100                 105                 110         


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
        115                 120                 125             


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
    130                 135                 140                 


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Asp Gly Thr Leu 
145                 150                 155                 160 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
                165                 170                 175     


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
            180                 185                 


<210>  21
<211>  653
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  21

Met Val Ser Lys Gly Glu Glu Val Ile Lys Glu Phe Met Arg Phe Lys 
1               5                   10                  15      


Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile Glu Gly 
            20                  25                  30          


Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys Leu Lys 
        35                  40                  45              


Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu Ser Pro 
    50                  55                  60                  


Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala Asp Ile 
65                  70                  75                  80  


Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp Glu Arg 
                85                  90                  95      


Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln Asp Ser 
            100                 105                 110         


Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg Gly Thr 
        115                 120                 125             


Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met Gly Trp 
    130                 135                 140                 


Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu Lys Gly 
145                 150                 155                 160 


Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr Leu Val 
                165                 170                 175     


Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu Pro Gly 
            180                 185                 190         


Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn Glu Asp 
        195                 200                 205             


Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His His Leu 
    210                 215                 220                 


Phe Leu Gly His Gly Thr Gly Ser Thr Gly Ser Gly Ser Ser Gly Thr 
225                 230                 235                 240 


Ala Ser Ser Glu Asp Asn Asn Met Ala Val Ile Lys Glu Phe Met Arg 
                245                 250                 255     


Phe Lys Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile 
            260                 265                 270         


Glu Gly Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys 
        275                 280                 285             


Leu Lys Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu 
    290                 295                 300                 


Ser Pro Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala 
305                 310                 315                 320 


Asp Ile Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp 
                325                 330                 335     


Glu Arg Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln 
            340                 345                 350         


Asp Ser Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg 
        355                 360                 365             


Gly Thr Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met 
    370                 375                 380                 


Gly Trp Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu 
385                 390                 395                 400 


Lys Gly Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr 
                405                 410                 415     


Leu Val Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu 
            420                 425                 430         


Pro Gly Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn 
        435                 440                 445             


Glu Asp Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His 
    450                 455                 460                 


His Leu Phe Leu Ser Gly Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
465                 470                 475                 480 


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
                485                 490                 495     


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
            500                 505                 510         


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
        515                 520                 525             


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
    530                 535                 540                 


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
545                 550                 555                 560 


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
                565                 570                 575     


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
            580                 585                 590         


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
        595                 600                 605             


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Asp Leu 
    610                 615                 620                 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
625                 630                 635                 640 


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
                645                 650             


<210>  22
<211>  653
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  22

Met Val Ser Lys Gly Glu Glu Val Ile Lys Glu Phe Met Arg Phe Lys 
1               5                   10                  15      


Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile Glu Gly 
            20                  25                  30          


Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys Leu Lys 
        35                  40                  45              


Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu Ser Pro 
    50                  55                  60                  


Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala Asp Ile 
65                  70                  75                  80  


Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp Glu Arg 
                85                  90                  95      


Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln Asp Ser 
            100                 105                 110         


Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg Gly Thr 
        115                 120                 125             


Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met Gly Trp 
    130                 135                 140                 


Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu Lys Gly 
145                 150                 155                 160 


Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr Leu Val 
                165                 170                 175     


Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu Pro Gly 
            180                 185                 190         


Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn Glu Asp 
        195                 200                 205             


Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His His Leu 
    210                 215                 220                 


Phe Leu Gly His Gly Thr Gly Ser Thr Gly Ser Gly Ser Ser Gly Thr 
225                 230                 235                 240 


Ala Ser Ser Glu Asp Asn Asn Met Ala Val Ile Lys Glu Phe Met Arg 
                245                 250                 255     


Phe Lys Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile 
            260                 265                 270         


Glu Gly Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys 
        275                 280                 285             


Leu Lys Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu 
    290                 295                 300                 


Ser Pro Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala 
305                 310                 315                 320 


Asp Ile Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp 
                325                 330                 335     


Glu Arg Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln 
            340                 345                 350         


Asp Ser Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg 
        355                 360                 365             


Gly Thr Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met 
    370                 375                 380                 


Gly Trp Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu 
385                 390                 395                 400 


Lys Gly Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr 
                405                 410                 415     


Leu Val Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu 
            420                 425                 430         


Pro Gly Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn 
        435                 440                 445             


Glu Asp Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His 
    450                 455                 460                 


His Leu Phe Leu Ser Gly Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
465                 470                 475                 480 


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
                485                 490                 495     


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
            500                 505                 510         


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
        515                 520                 525             


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
    530                 535                 540                 


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
545                 550                 555                 560 


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
                565                 570                 575     


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
            580                 585                 590         


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
        595                 600                 605             


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Thr Leu 
    610                 615                 620                 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
625                 630                 635                 640 


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
                645                 650             


<210>  23
<211>  653
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  23

Met Val Ser Lys Gly Glu Glu Val Ile Lys Glu Phe Met Arg Phe Lys 
1               5                   10                  15      


Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile Glu Gly 
            20                  25                  30          


Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys Leu Lys 
        35                  40                  45              


Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu Ser Pro 
    50                  55                  60                  


Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala Asp Ile 
65                  70                  75                  80  


Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp Glu Arg 
                85                  90                  95      


Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln Asp Ser 
            100                 105                 110         


Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg Gly Thr 
        115                 120                 125             


Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met Gly Trp 
    130                 135                 140                 


Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu Lys Gly 
145                 150                 155                 160 


Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr Leu Val 
                165                 170                 175     


Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu Pro Gly 
            180                 185                 190         


Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn Glu Asp 
        195                 200                 205             


Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His His Leu 
    210                 215                 220                 


Phe Leu Gly His Gly Thr Gly Ser Thr Gly Ser Gly Ser Ser Gly Thr 
225                 230                 235                 240 


Ala Ser Ser Glu Asp Asn Asn Met Ala Val Ile Lys Glu Phe Met Arg 
                245                 250                 255     


Phe Lys Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile 
            260                 265                 270         


Glu Gly Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys 
        275                 280                 285             


Leu Lys Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu 
    290                 295                 300                 


Ser Pro Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala 
305                 310                 315                 320 


Asp Ile Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp 
                325                 330                 335     


Glu Arg Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln 
            340                 345                 350         


Asp Ser Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg 
        355                 360                 365             


Gly Thr Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met 
    370                 375                 380                 


Gly Trp Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu 
385                 390                 395                 400 


Lys Gly Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr 
                405                 410                 415     


Leu Val Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu 
            420                 425                 430         


Pro Gly Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn 
        435                 440                 445             


Glu Asp Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His 
    450                 455                 460                 


His Leu Phe Leu Ser Gly Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
465                 470                 475                 480 


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
                485                 490                 495     


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
            500                 505                 510         


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
        515                 520                 525             


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
    530                 535                 540                 


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
545                 550                 555                 560 


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
                565                 570                 575     


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
            580                 585                 590         


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
        595                 600                 605             


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Asp Leu 
    610                 615                 620                 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
625                 630                 635                 640 


Asp Pro Thr Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
                645                 650             


<210>  24
<211>  653
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  24

Met Val Ser Lys Gly Glu Glu Val Ile Lys Glu Phe Met Arg Phe Lys 
1               5                   10                  15      


Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile Glu Gly 
            20                  25                  30          


Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys Leu Lys 
        35                  40                  45              


Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu Ser Pro 
    50                  55                  60                  


Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala Asp Ile 
65                  70                  75                  80  


Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp Glu Arg 
                85                  90                  95      


Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln Asp Ser 
            100                 105                 110         


Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg Gly Thr 
        115                 120                 125             


Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met Gly Trp 
    130                 135                 140                 


Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu Lys Gly 
145                 150                 155                 160 


Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr Leu Val 
                165                 170                 175     


Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu Pro Gly 
            180                 185                 190         


Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn Glu Asp 
        195                 200                 205             


Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His His Leu 
    210                 215                 220                 


Phe Leu Gly His Gly Thr Gly Ser Thr Gly Ser Gly Ser Ser Gly Thr 
225                 230                 235                 240 


Ala Ser Ser Glu Asp Asn Asn Met Ala Val Ile Lys Glu Phe Met Arg 
                245                 250                 255     


Phe Lys Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile 
            260                 265                 270         


Glu Gly Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys 
        275                 280                 285             


Leu Lys Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu 
    290                 295                 300                 


Ser Pro Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala 
305                 310                 315                 320 


Asp Ile Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp 
                325                 330                 335     


Glu Arg Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln 
            340                 345                 350         


Asp Ser Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg 
        355                 360                 365             


Gly Thr Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met 
    370                 375                 380                 


Gly Trp Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu 
385                 390                 395                 400 


Lys Gly Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr 
                405                 410                 415     


Leu Val Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu 
            420                 425                 430         


Pro Gly Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn 
        435                 440                 445             


Glu Asp Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His 
    450                 455                 460                 


His Leu Phe Leu Ser Gly Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
465                 470                 475                 480 


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
                485                 490                 495     


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
            500                 505                 510         


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
        515                 520                 525             


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
    530                 535                 540                 


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
545                 550                 555                 560 


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
                565                 570                 575     


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Ala Ile Thr Leu Asp Glu 
            580                 585                 590         


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
        595                 600                 605             


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Thr Leu 
    610                 615                 620                 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
625                 630                 635                 640 


Asp Pro Thr Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
                645                 650             


<210>  25
<211>  653
<212>  PRT
<213>  artificial

<220>
<223>  mutant aequorin

<400>  25

Met Val Ser Lys Gly Glu Glu Val Ile Lys Glu Phe Met Arg Phe Lys 
1               5                   10                  15      


Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile Glu Gly 
            20                  25                  30          


Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys Leu Lys 
        35                  40                  45              


Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu Ser Pro 
    50                  55                  60                  


Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala Asp Ile 
65                  70                  75                  80  


Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp Glu Arg 
                85                  90                  95      


Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln Asp Ser 
            100                 105                 110         


Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg Gly Thr 
        115                 120                 125             


Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met Gly Trp 
    130                 135                 140                 


Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu Lys Gly 
145                 150                 155                 160 


Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr Leu Val 
                165                 170                 175     


Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu Pro Gly 
            180                 185                 190         


Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn Glu Asp 
        195                 200                 205             


Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His His Leu 
    210                 215                 220                 


Phe Leu Gly His Gly Thr Gly Ser Thr Gly Ser Gly Ser Ser Gly Thr 
225                 230                 235                 240 


Ala Ser Ser Glu Asp Asn Asn Met Ala Val Ile Lys Glu Phe Met Arg 
                245                 250                 255     


Phe Lys Val Arg Met Glu Gly Ser Met Asn Gly His Glu Phe Glu Ile 
            260                 265                 270         


Glu Gly Glu Gly Glu Gly Arg Pro Tyr Glu Gly Thr Gln Thr Ala Lys 
        275                 280                 285             


Leu Lys Val Thr Lys Gly Gly Pro Leu Pro Phe Ala Trp Asp Ile Leu 
    290                 295                 300                 


Ser Pro Gln Phe Met Tyr Gly Ser Lys Ala Tyr Val Lys His Pro Ala 
305                 310                 315                 320 


Asp Ile Pro Asp Tyr Lys Lys Leu Ser Phe Pro Glu Gly Phe Lys Trp 
                325                 330                 335     


Glu Arg Val Met Asn Phe Glu Asp Gly Gly Leu Val Thr Val Thr Gln 
            340                 345                 350         


Asp Ser Ser Leu Gln Asp Gly Thr Leu Ile Tyr Lys Val Lys Met Arg 
        355                 360                 365             


Gly Thr Asn Phe Pro Pro Asp Gly Pro Val Met Gln Lys Lys Thr Met 
    370                 375                 380                 


Gly Trp Glu Ala Ser Thr Glu Arg Leu Tyr Pro Arg Asp Gly Val Leu 
385                 390                 395                 400 


Lys Gly Glu Ile His Gln Ala Leu Lys Leu Lys Asp Gly Gly His Tyr 
                405                 410                 415     


Leu Val Glu Phe Lys Thr Ile Tyr Met Ala Lys Lys Pro Val Gln Leu 
            420                 425                 430         


Pro Gly Tyr Tyr Tyr Val Asp Thr Lys Leu Asp Ile Thr Ser His Asn 
        435                 440                 445             


Glu Asp Tyr Thr Ile Val Glu Gln Tyr Glu Arg Ser Glu Gly Arg His 
    450                 455                 460                 


His Leu Phe Leu Ser Gly Phe Asp Asn Pro Arg Trp Ile Gly Arg His 
465                 470                 475                 480 


Lys His Met Phe Asn Phe Leu Asp Val Asn His Asn Gly Lys Ile Ser 
                485                 490                 495     


Leu Asp Glu Met Val Tyr Lys Ala Ser Asp Ile Val Ile Asn Asn Leu 
            500                 505                 510         


Gly Ala Thr Pro Glu Gln Ala Lys Arg His Lys Asp Ala Val Glu Ala 
        515                 520                 525             


Phe Phe Gly Gly Ala Gly Met Lys Tyr Gly Val Glu Thr Asp Trp Pro 
    530                 535                 540                 


Ala Tyr Ile Glu Gly Trp Lys Lys Leu Ala Thr Asp Glu Leu Glu Lys 
545                 550                 555                 560 


Tyr Ala Lys Asn Glu Pro Thr Leu Ile Arg Ile Trp Gly Asp Ala Leu 
                565                 570                 575     


Phe Asp Ile Val Asp Lys Asp Gln Asn Gly Asp Ile Thr Leu Asp Glu 
            580                 585                 590         


Trp Lys Ala Tyr Thr Lys Ala Ala Gly Ile Ile Gln Ser Ser Glu Asp 
        595                 600                 605             


Cys Glu Glu Thr Phe Arg Val Cys Asp Ile Asp Glu Ser Gly Thr Leu 
    610                 615                 620                 


Asp Val Asp Glu Met Thr Arg Gln His Leu Gly Phe Trp Tyr Thr Met 
625                 630                 635                 640 


Asp Pro Ala Cys Glu Lys Leu Tyr Gly Gly Ala Val Pro 
                645                 650             


