                         SEQUENCE LISTING

<110>  VIB VZW
       UNIVERSITEIT GENT
       THE UNIVERSITY OF WESTERN ONTARIO
 
<120>  NOVEL CLASS OF ANTIFREEZE PROTEINS

<130>  TVL/ANTI/408

<150>  EP 13189511.2
<151>  2013-10-21

<160>  27    

<170>  PatentIn version 3.5

<210>  1
<211>  9
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Antifreeze protein


<220>
<221>  MISC_FEATURE
<222>  (5)..(5)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  MISC_FEATURE
<222>  (8)..(8)
<223>  Xaa can be any naturally occurring amino acid

<400>  1

Cys Thr Asn Cys Xaa Asn Cys Xaa Asn 
1               5                   


<210>  2
<211>  13
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Antifreeze protein


<220>
<221>  MISC_FEATURE
<222>  (5)..(5)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  MISC_FEATURE
<222>  (8)..(8)
<223>  Xaa can be any naturally occurring amino acid

<400>  2

Cys Thr Asn Cys Xaa Asn Cys Xaa Asn Cys Thr Asn Cys 
1               5                   10              


<210>  3
<211>  27
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Antifreeze protein


<220>
<221>  MISC_FEATURE
<222>  (2)..(2)
<223>  Xaa can be Val or Ile

<220>
<221>  MISC_FEATURE
<222>  (5)..(5)
<223>  Xaa can be Val or Thr

<220>
<221>  MISC_FEATURE
<222>  (6)..(6)
<223>  Xaa can be Gly or Asn

<220>
<221>  MISC_FEATURE
<222>  (11)..(11)
<223>  Xaa can be Ser, Ala, Ile, Val or Thr

<220>
<221>  MISC_FEATURE
<222>  (14)..(14)
<223>  Xaa can be Ile, Val, Ser or Thr

<220>
<221>  MISC_FEATURE
<222>  (21)..(21)
<223>  Xaa can be Asp, Gly or Asn

<220>
<221>  MISC_FEATURE
<222>  (23)..(23)
<223>  Xaa can be any naturally occuring amino acid

<220>
<221>  MISC_FEATURE
<222>  (24)..(24)
<223>  Xaa can be Asn, Ser or Arg

<220>
<221>  MISC_FEATURE
<222>  (26)..(26)
<223>  Xaa can be any naturally occurring amino acid

<400>  3

Cys Xaa Asn Cys Xaa Xaa Cys Thr Asn Cys Xaa Asn Cys Xaa Asn Cys 
1               5                   10                  15      


Thr Asn Cys His Xaa Cys Xaa Xaa Cys Xaa Asp 
            20                  25          


<210>  4
<211>  141
<212>  PRT
<213>  Tetranychus urticae


<220>
<221>  SIGNAL
<222>  (1)..(24)

<400>  4

Met His Leu Lys Thr His Ile Leu Ile Leu Val Ser Val Phe Met Ile 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Thr Asp Ser Asn Gly Asn Val 
            20                  25                  30          


Asn Cys Val Asn Cys Thr Gly Cys Thr Asn Cys Val Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Glu Ser Cys Ser Asp Ser Thr Asn Cys 
    50                  55                  60                  


Arg Ser Ser Arg Asn Cys Thr Ser Cys Thr Asp Cys Ser Asp Cys Lys 
65                  70                  75                  80  


Asn Cys Thr Ser Cys Thr Asp Ala Lys Asn Cys Asn Asp Ser Arg Ser 
                85                  90                  95      


Cys Thr Asn Cys Arg Asn Cys Ser Asn Cys Ala Ser Cys Thr Asn Cys 
            100                 105                 110         


Arg Asp Cys Gln Arg Cys Thr His Cys Thr Asn Val Arg Asn Ser Ala 
        115                 120                 125             


Asn Cys Met Asn Arg Thr Asp Cys Thr Asn Cys Lys Cys 
    130                 135                 140     


<210>  5
<211>  123
<212>  PRT
<213>  Tetranychus urticae

<400>  5

Met Tyr Leu Lys Thr Tyr Phe Leu Ile Leu Val Ser Ala Gly Met Val 
1               5                   10                  15      


Ser Leu Met Thr Pro Thr Val Asp Gly Asp Gly Asn Thr Asn Cys Val 
            20                  25                  30          


Asn Cys Val Gly Cys Thr Asn Cys Ser Asn Cys Ile Asn Cys Thr Asn 
        35                  40                  45              


Cys His Asp Cys Ala Asn Cys Thr Asp Ser Glu Asn Cys Ser Asn Ser 
    50                  55                  60                  


Arg Asn Thr Ala Asn Ser Asn Gly Cys Gln Asn Cys Ala Asn Cys Thr 
65                  70                  75                  80  


Tyr Cys Val Ser Cys Ser Asp Cys Ala Asp Cys Ala Ser Ser Lys Gly 
                85                  90                  95      


Cys Ala Asn Cys Ala Asn Cys Thr Asn Cys Thr Asn Cys Arg Asn Cys 
            100                 105                 110         


Ala Asn Arg Ala Asn Leu Ser Asp Thr Gln Cys 
        115                 120             


<210>  6
<211>  210
<212>  PRT
<213>  Tetranychus urticae

<400>  6

Met Tyr Leu Lys Ser His Ile Phe Ile Leu Val Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Asp Gly Ser Asp Gly Asn Gln 
            20                  25                  30          


Asn Cys Ile Asn Cys Val Gly Cys Thr Asn Cys Ala Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Gln Asn Cys Ala Asp Cys Thr Asn Gly 
    50                  55                  60                  


Thr Ser Leu Gln Asn Cys Lys Asn Ser Thr Asp Cys Asn Arg Cys Ala 
65                  70                  75                  80  


Asn Cys Ile Tyr Cys Leu Tyr Cys Thr Asp Cys Val Gly Cys Thr Asn 
                85                  90                  95      


Cys Arg Asn Cys Ser Asn Ser Ser Asn Cys Thr Asp Ser Thr Asp Gly 
            100                 105                 110         


Ser Tyr Leu Gln Asn Cys Lys Asn Cys Lys Asp Cys Lys Arg Cys Ser 
        115                 120                 125             


Asn Cys Ala Asn Cys Leu Arg Cys Lys Asp Cys Ser Asn Cys Thr Asp 
    130                 135                 140                 


Cys Val Tyr Ser Ser Asp Cys Gln Asn Cys Ile Asp Cys Thr Asn Cys 
145                 150                 155                 160 


Ser Tyr Leu Gln Asn Cys Lys Thr Cys Thr Asp Cys Lys Arg Cys Asn 
                165                 170                 175     


Asn Cys Gln Asp Cys Leu Arg Cys Ser Gly Cys Asn Asn Cys Lys Asn 
            180                 185                 190         


Cys Val Asp Ser Ala Asn Cys Asn Asp Arg Thr Asp Cys Lys Asn Cys 
        195                 200                 205             


His Cys 
    210 


<210>  7
<211>  92
<212>  PRT
<213>  Tetranychus urticae

<400>  7

Met Thr Pro Val Asp Ser Tyr Gly Asn Thr Asn Cys Lys Asn Cys Val 
1               5                   10                  15      


Asn Cys Thr Asp Cys Ala Asp Cys Gln Asp Cys Glu Asn Cys Ala Asn 
            20                  25                  30          


Cys Thr Gly Ser Lys Asn Cys Thr Asn Cys Val Asn Cys Ser Asn Cys 
        35                  40                  45              


Asp Asp Cys Ala Asn Cys Ser Gly Ser Lys Asn Cys Ser His Cys Asp 
    50                  55                  60                  


Asn Cys Thr Arg Cys Asn Asp Cys Glu Lys Cys Asn Thr Arg Lys Asp 
65                  70                  75                  80  


Cys Ala His Arg Thr Ser Leu Thr Asp Cys Gln Cys 
                85                  90          


<210>  8
<211>  153
<212>  PRT
<213>  Tetranychus urticae

<400>  8

Met Tyr Leu Lys Thr His Phe Phe Ile Leu Ala Ser Leu Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Thr Asp Ser Asp Gly Asn Thr 
            20                  25                  30          


Asn Cys Val Asn Cys Thr Gly Cys Thr Asn Cys Ser Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Glu Ser Cys Ser Asp Ser Thr Asn Cys 
    50                  55                  60                  


Arg Asn Ser Arg Asn Cys Ala Ser Cys Thr Asp Cys Ser Asp Cys Lys 
65                  70                  75                  80  


Asn Cys Ala Ser Cys Thr Ala Ser Lys Asn Cys Asn Asp Cys Arg Ser 
                85                  90                  95      


Cys Thr Ser Cys Thr Asn Cys Ser Asn Cys Ala Asn Cys Thr Asn Ser 
            100                 105                 110         


Arg Asn Cys Ser Asp Cys Thr Asn Cys Thr Lys Cys Thr Asp Cys Gln 
        115                 120                 125             


Arg Cys Thr Asn Cys Thr Asn Val Lys Asn Ser Ala Asn Cys Leu Asn 
    130                 135                 140                 


Arg Thr Asp Cys Thr Asn Cys Gln Cys 
145                 150             


<210>  9
<211>  159
<212>  PRT
<213>  Tetranychus urticae

<400>  9

Met Tyr Leu Lys Thr His Ile Phe Ile Leu Val Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Asp Gly Asn Gln Asn Cys Ile 
            20                  25                  30          


Asn Cys Val Gly Cys Thr Asn Cys Ala Asn Cys Ile Asn Cys Thr Asn 
        35                  40                  45              


Cys His Asn Cys Ala Asn Cys Asn Asp Cys Thr Asn Cys Lys Asp Cys 
    50                  55                  60                  


Arg Asn Cys Asp Asn Cys Arg Asp Cys Gln Arg Cys Ala Asp Cys Thr 
65                  70                  75                  80  


Lys Cys Gln Gly Cys Asp Asp Thr Val Gly Ser Lys Lys Cys Arg Asp 
                85                  90                  95      


Cys Ser Asp Cys Ser Asn Cys Arg Asp Cys Ile Asn Cys Ser Arg Cys 
            100                 105                 110         


Arg Asn Cys Glu Ser Cys Thr Asp Cys Gln Arg Cys Ala Asn Cys Ala 
        115                 120                 125             


Asn Cys Leu Arg Cys Ser Asp Cys Arg Asp Cys Thr Arg Cys Thr Asp 
    130                 135                 140                 


Ser Gln Arg Cys Ala Asp Arg Thr Asp Cys Lys Asn Cys Gln Cys 
145                 150                 155                 


<210>  10
<211>  209
<212>  PRT
<213>  Tetranychus urticae

<400>  10

Met Tyr Leu Lys Thr His Ile Phe Ile Leu Leu Ser Ile Phe Met Val 
1               5                   10                  15      


Ser Leu Thr Thr Pro Ser Thr Gly Ser Gly Ser Asp Gly Asn Gln Asn 
            20                  25                  30          


Cys Val Asn Cys Val Asn Cys Thr Asn Cys Ile Asn Cys Thr Asn Cys 
        35                  40                  45              


Thr Asn Cys His Gly Cys Gln Asn Cys Glu Asp Cys Thr Asn Gly Ile 
    50                  55                  60                  


Ser Leu Arg Tyr Cys Lys Asn Ser Arg Asp Cys Asn Arg Cys Asp Val 
65                  70                  75                  80  


Cys Thr Cys Cys Phe Asn Cys Thr Asp Cys Val Tyr Cys Thr Asn Cys 
                85                  90                  95      


Arg Asn Cys Ser Asn Ser Ser Asn Cys Ile Asp Ser Thr Asp Gly Ser 
            100                 105                 110         


Tyr Leu Arg Tyr Cys Lys Thr Cys Lys Asp Cys Lys Arg Cys Asp Asn 
        115                 120                 125             


Cys Asn Asn Cys Leu Arg Cys Lys Asp Cys Ile Tyr Cys Thr Asp Cys 
    130                 135                 140                 


Val Tyr Cys Ser Asp Ser Ser Asn Cys Glu Asp Cys Thr Asn Gly Ser 
145                 150                 155                 160 


Tyr Leu Gln Asn Cys Lys Thr Cys Thr Asp Cys Lys Arg Cys Ser Gly 
                165                 170                 175     


Cys Lys Asp Cys Leu Arg Cys Ser Asn Cys Ala Asn Cys Glu Asn Cys 
            180                 185                 190         


Val Asp Ala Ala Asn Cys Asn Asn Arg Thr Asp Cys Thr Asn Cys Gln 
        195                 200                 205             


Cys 
    


<210>  11
<211>  135
<212>  PRT
<213>  Tetranychus urticae

<400>  11

Met Tyr Val Lys Thr Tyr Phe Leu Ile Leu Val Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Asp Gly Asn Thr Asn Cys Val 
            20                  25                  30          


Asn Cys Val Gly Cys Thr Asn Cys Ala Asn Cys Ile Asn Cys Thr Asn 
        35                  40                  45              


Cys His Asp Cys Ala Asn Cys Thr Asp Ser Thr Asn Cys Ser Asn Ser 
    50                  55                  60                  


Arg Asn Cys Glu Ser Ser Ile Asp Cys Gln Arg Cys Ala Asn Gly Ile 
65                  70                  75                  80  


Asn Cys Val Gly Cys Ser Asp Cys Ala Asp Cys Thr Gly Cys Arg Asn 
                85                  90                  95      


Cys Ser Asn Cys Ala Asn Cys Ser Gln Cys Thr Gly Cys Gln Arg Cys 
            100                 105                 110         


Ala Asn Cys Thr Asn Cys Thr Asn Cys Lys Asp Cys Ala Asn Arg Thr 
        115                 120                 125             


Gly Leu Thr Asn Cys Gln Cys 
    130                 135 


<210>  12
<211>  153
<212>  PRT
<213>  Tetranychus urticae

<400>  12

Met Phe Ala Lys Thr Tyr Phe Phe Ile Leu Val Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Leu Ser Ala Gly Ser Thr Asp Ser Asp Gly Asn Thr 
            20                  25                  30          


Asn Cys Val Asn Cys Thr Gly Cys Thr Asn Cys Ser Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Glu Ser Cys Ser Asp Ser Thr Asn Cys 
    50                  55                  60                  


His Asn Ser Ile Asn Cys Ala Asn Cys Thr Asp Cys Lys Asp Cys Lys 
65                  70                  75                  80  


Asn Cys Asn Asp Cys Thr Ala Ser Gly Asn Cys Glu Gly Ser Arg Asn 
                85                  90                  95      


Cys Thr Thr Cys Thr Tyr Cys Ser Asn Cys Ala Asp Cys Thr Ser Ser 
            100                 105                 110         


Arg Asn Cys Ser Asp Cys Ala Asn Cys Thr Ala Cys Lys Asp Cys Gln 
        115                 120                 125             


Gly Cys Ser Asn Cys Thr Thr Cys Thr Asn Ser Ser Asn Cys Lys Asn 
    130                 135                 140                 


Arg Thr Gly Cys Thr Asn Cys Gln Cys 
145                 150             


<210>  13
<211>  168
<212>  PRT
<213>  Tetranychus urticae

<400>  13

Met Tyr Leu Lys Thr His Ile Leu Ile Leu Val Ser Ile Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Gly Gly Ser Asp Gly Ser Pro 
            20                  25                  30          


Asp Ser Asp Gly Asn Ile Asn Cys Val Asn Cys Val Gly Cys Thr Asn 
        35                  40                  45              


Cys Ala Asn Cys Val Asn Cys Thr Asn Cys His Gly Cys Lys Asn Cys 
    50                  55                  60                  


Asn Asp Cys Thr Asn Cys Thr Asn Cys Lys Asn Cys Asp Asn Cys Arg 
65                  70                  75                  80  


Asp Cys Gln Arg Cys Thr Asn Cys Pro Asn Cys Leu Ser Cys Glu Asp 
                85                  90                  95      


Cys Ser Asp Cys Lys Asn Leu Arg Asn Cys Ser Asn Cys Ala Asn Cys 
            100                 105                 110         


Arg Glu Cys Ile Asn Cys Ser Gly Cys Lys Asn Cys Glu Asn Cys Thr 
        115                 120                 125             


Asp Cys Lys Arg Cys Ser Asn Cys Ser Asp Cys Leu Arg Cys Thr Asp 
    130                 135                 140                 


Cys Lys Asn Cys Thr Ser Cys Glu Asp Ser Ala Asn Cys Asn Asn Arg 
145                 150                 155                 160 


Thr Asp Cys Lys Asn Cys Gln Cys 
                165             


<210>  14
<211>  141
<212>  PRT
<213>  Tetranychus urticae

<400>  14

Met His Leu Lys Thr His Ile Leu Ile Leu Val Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Ala Asp Ser Asn Gly Asn Val 
            20                  25                  30          


Asn Cys Val Asn Cys Thr Gly Cys Thr Asn Cys Val Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Glu Ser Cys Ser Asp Ser Thr Arg Cys 
    50                  55                  60                  


Arg Asn Ser Arg Asn Cys Ala Ser Cys Thr Asp Cys Ser Asp Cys Lys 
65                  70                  75                  80  


Asn Cys Thr Ser Cys Thr Asp Ser Lys Asn Cys Asn Asp Ser Arg Ser 
                85                  90                  95      


Cys Thr Asn Cys Lys Asn Cys Ser Asn Cys Ala Ser Cys Thr Asn Cys 
            100                 105                 110         


Ala Asp Cys Gln Arg Cys Thr Asn Cys Thr Asn Val Lys Asn Ser Ala 
        115                 120                 125             


Asp Cys Met Asp Arg Thr Gly Cys Thr Asn Cys Lys Cys 
    130                 135                 140     


<210>  15
<211>  153
<212>  PRT
<213>  Tetranychus urticae

<400>  15

Met Tyr Leu Lys Thr His Ile Phe Ile Leu Ile Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ser Ile Pro Ser Thr Gly Ser Ala Asp Ser Asn Gly Asn Val 
            20                  25                  30          


Asn Cys Val Asn Cys Thr Gly Cys Thr Asn Cys Val Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Glu Ser Cys Ser Asp Ser Thr Arg Cys 
    50                  55                  60                  


Arg Asn Ser Arg Asn Cys Ala Ser Cys Thr Asp Cys Ser Asp Cys Lys 
65                  70                  75                  80  


Asn Cys Ala Ser Cys Thr Ala Ser Lys Asn Cys Asn Asp Ser Arg Ser 
                85                  90                  95      


Cys Thr Asn Cys Thr Asn Cys Ser Asn Cys Thr Asn Cys Ile Asn Ser 
            100                 105                 110         


Arg Asn Cys Ser Asp Cys Thr Asn Cys Thr Lys Cys Thr Asp Cys Gln 
        115                 120                 125             


Arg Cys Thr Asn Cys Thr Asn Val Lys Asn Ser Ala Asn Cys Thr Asn 
    130                 135                 140                 


Arg Thr Gly Cys Thr Asn Cys Lys Cys 
145                 150             


<210>  16
<211>  128
<212>  PRT
<213>  Tetranychus urticae

<400>  16

Met Tyr Leu Lys Thr His Ile Phe Ile Leu Ile Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Phe Thr Gly Ser Thr Asp Ser Asp Gly Asn Gln 
            20                  25                  30          


Asn Cys Val Asn Cys Asn Cys Val Asn Cys Thr Asn Cys His Gly Cys 
        35                  40                  45              


Gln Asn Cys Lys Asp Cys Thr Asp Cys Thr Asn Cys Asn Asp Cys Arg 
    50                  55                  60                  


Asp Cys Gln Arg Cys Ala Asn Ser Thr Asp Cys Leu Ser Cys Gly Asp 
65                  70                  75                  80  


Gly Ala Asp Cys Lys Asn Cys Arg Asn Cys Ser Asn Cys Ala Asn Cys 
                85                  90                  95      


Arg Asp Cys Ile Asp Cys Ser Asn Cys Ala Asn Cys Asn Arg Ile Thr 
            100                 105                 110         


Asp Ser Lys Gly Cys Ala Asp Arg Thr Asp Cys Thr Gln Cys Gln Cys 
        115                 120                 125             


<210>  17
<211>  95
<212>  PRT
<213>  Tetranychus urticae

<400>  17

Met Thr Ser Ile Asp Ser Tyr Gly Asn Thr Asn Cys Lys Asn Cys Val 
1               5                   10                  15      


Asn Cys Thr Asp Cys Thr Asp Cys Gln Asp Cys Asn Asp Cys Lys Asn 
            20                  25                  30          


Cys Ala Asn Cys Thr Gly Ser Arg Asn Cys Thr Asn Cys Val Asn Cys 
        35                  40                  45              


Ser Asn Cys Glu Asp Cys Ala Asn Cys Thr Gly Ser Lys Asn Cys Val 
    50                  55                  60                  


Asp Cys Ala Asn Cys Ile Gln Cys Thr Asn Cys Gln Lys Cys Ala Asn 
65                  70                  75                  80  


Arg Val Asn Cys Ala Asn Arg Thr Gly Leu Thr Asn Cys Gln Cys 
                85                  90                  95  


<210>  18
<211>  160
<212>  PRT
<213>  Tetranychus urticae

<400>  18

Met Tyr Leu Lys Thr His Phe Phe Leu Leu Ala Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Val Thr Pro Ser Thr Gly Ser Gly Gly Ser His Gly Ser Pro 
            20                  25                  30          


Asp Ser Asp Gly Asn Ile Asn Cys Val Asn Cys Lys Gly Cys Thr Asn 
        35                  40                  45              


Cys Thr Asn Cys Ile Asn Cys Ile Asn Cys His Gly Cys Glu Asp Cys 
    50                  55                  60                  


Ser Asp Cys Thr Arg Cys Val Asn Cys Ser Gly Asn Ser Ser Ile Leu 
65                  70                  75                  80  


Thr Asp Cys Ser Asn Cys Val Asn Cys Ser Ser Cys Thr Asn Cys Ser 
                85                  90                  95      


Asp Cys Lys Asn Ser Ser Ser Cys Thr Asp Ser Lys Asn Cys Asn Asp 
            100                 105                 110         


Ser Ser Ser Leu Thr Asn Cys Thr Asn Cys Ser Asn Cys Thr Ser Cys 
        115                 120                 125             


Thr Lys Cys Thr Asp Cys Gln Asn Cys Asp Ser Cys Thr Asn Val Gln 
    130                 135                 140                 


Asn Ser Ala Asn Cys Lys Ser Arg Thr Gly Cys Ser His Cys Gln Cys 
145                 150                 155                 160 


<210>  19
<211>  141
<212>  PRT
<213>  Tetranychus urticae

<400>  19

Met His Leu Lys Thr His Ile Leu Ile Leu Val Ser Val Phe Met Ile 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Thr Asp Ser Asn Gly Asn Val 
            20                  25                  30          


Asn Cys Val Asn Cys Thr Gly Cys Thr Asn Cys Val Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Glu Arg Cys Ser Asp Ser Thr Asn Cys 
    50                  55                  60                  


Arg Ser Ser Arg Asn Cys Thr Ser Cys Thr Asp Cys Ser Asp Cys Lys 
65                  70                  75                  80  


Asn Cys Thr Ser Cys Thr Asp Ala Lys Asn Cys Asn Asp Ser Arg Ser 
                85                  90                  95      


Cys Thr Asn Cys Arg Asn Cys Ser Asn Cys Ala Ser Cys Thr Asn Cys 
            100                 105                 110         


Arg Asp Cys Gln Arg Cys Thr Asn Cys Thr Asn Val Arg Asn Ser Ala 
        115                 120                 125             


Asn Cys Met Asn Arg Thr Asp Cys Thr Asn Cys Lys Cys 
    130                 135                 140     


<210>  20
<211>  123
<212>  PRT
<213>  Tetranychus urticae

<400>  20

Met Tyr Leu Lys Thr Tyr Phe Leu Ile Leu Val Ser Ala Gly Met Val 
1               5                   10                  15      


Ser Leu Met Thr Pro Thr Val Asp Gly Asp Gly Asn Thr Asn Cys Val 
            20                  25                  30          


Asn Cys Val Gly Cys Thr Asn Cys Ser Asn Cys Ile Asn Cys Thr Asn 
        35                  40                  45              


Cys His Asp Cys Ala Asn Cys Thr Asp Ser Glu Asn Cys Ser Asn Ser 
    50                  55                  60                  


Arg Asn Thr Ala Asn Ser Asn Gly Cys Gln Asn Cys Ala Asn Cys Thr 
65                  70                  75                  80  


Tyr Cys Val Ser Cys Ser Asp Cys Ala Asp Cys Ala Ser Ser Lys Gly 
                85                  90                  95      


Cys Ala Asn Cys Ala Asn Cys Thr Asn Cys Ile Asn Cys Arg Asn Cys 
            100                 105                 110         


Ala Asn Arg Ala Asn Leu Ser Asp Thr Gln Cys 
        115                 120             


<210>  21
<211>  210
<212>  PRT
<213>  Tetranychus urticae

<400>  21

Met Tyr Leu Lys Ser His Ile Phe Ile Leu Val Ser Val Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Thr Asp Gly Ser Asp Gly Asn Gln 
            20                  25                  30          


Asn Cys Ile Asn Cys Val Gly Cys Thr Asn Cys Ala Asn Cys Ile Asn 
        35                  40                  45              


Cys Thr Asn Cys His Gly Cys Gln Asn Cys Ala Asp Cys Thr Asn Gly 
    50                  55                  60                  


Thr Ser Leu Gln Asn Cys Lys Asn Ser Thr Asp Cys Asn Arg Cys Ala 
65                  70                  75                  80  


Asn Cys Thr Tyr Cys Leu Tyr Cys Thr Asp Cys Val Gly Cys Thr Asn 
                85                  90                  95      


Cys Arg Asn Cys Ser Asn Ser Ser Asn Cys Thr Asp Ser Thr Asp Gly 
            100                 105                 110         


Ser Tyr Leu Gln Asn Cys Lys Asn Cys Lys Asp Cys Lys Arg Cys Ser 
        115                 120                 125             


Asn Cys Ala Asn Cys Leu Arg Cys Lys Asp Cys Ser Asn Cys Thr Asp 
    130                 135                 140                 


Cys Val Tyr Ser Ser Asp Cys Gln Asn Cys Ile Asp Cys Thr Asn Cys 
145                 150                 155                 160 


Ser Tyr Ser Gln Asn Cys Lys Thr Cys Thr Asp Cys Glu Arg Cys Asn 
                165                 170                 175     


Asn Cys Gln Asp Cys Leu Arg Cys Ser Gly Cys Asn Asn Cys Lys Lys 
            180                 185                 190         


Cys Val Asp Ser Glu Asn Cys Asn Asn Arg Thr Asp Cys Lys Asn Cys 
        195                 200                 205             


His Cys 
    210 


<210>  22
<211>  159
<212>  PRT
<213>  Tetranychus urticae

<400>  22

Met Tyr Leu Lys Thr His Ile Phe Ile Leu Val Ser Ile Phe Met Val 
1               5                   10                  15      


Ser Leu Ile Thr Pro Ser Thr Gly Ser Asp Gly Asn Gln Asn Cys Ile 
            20                  25                  30          


Asn Cys Val Gly Cys Thr Asn Cys Ala Asn Cys Ile Asn Cys Thr Asn 
        35                  40                  45              


Cys His Asn Cys Ala Asn Cys Asn Asp Cys Thr Asn Cys Lys Asp Cys 
    50                  55                  60                  


Arg Asn Cys Asp Asp Cys Arg Asp Cys Ser Arg Cys Ala Asp Cys Thr 
65                  70                  75                  80  


Lys Cys Glu Gly Cys Asp Asp Thr Val Gly Ser Lys Lys Cys Arg Asp 
                85                  90                  95      


Cys Ser Asp Cys Ser Asn Cys Arg Asp Cys Ile Asn Cys Ser Arg Cys 
            100                 105                 110         


Arg Asn Cys Glu Ser Cys Ala Asp Cys Gln Arg Cys Ala Asn Cys Ala 
        115                 120                 125             


Asn Cys Leu Arg Cys Ser Asp Cys Arg Asp Cys Thr Arg Cys Thr Asp 
    130                 135                 140                 


Ser Gln Arg Cys Ala Asp Arg Thr Asp Cys Lys Asn Cys Gln Cys 
145                 150                 155                 


<210>  23
<211>  92
<212>  PRT
<213>  Tetranychus urticae

<400>  23

Met Thr Pro Val Asp Ser Tyr Gly Asn Thr Asn Cys Lys Asn Cys Val 
1               5                   10                  15      


Asn Cys Thr Asp Cys Ala Asp Cys Gln Asp Cys Glu Asn Cys Ala Asn 
            20                  25                  30          


Cys Thr Gly Ser Lys Asn Cys Thr Asn Cys Val Asn Cys Ser Asn Cys 
        35                  40                  45              


Asp Asp Cys Ala Asn Cys Ser Gly Ser Lys Asn Cys Ser His Cys Asp 
    50                  55                  60                  


Asn Cys Thr Arg Cys Asn Asp Cys Glu Lys Cys Asn Thr Arg Lys Asp 
65                  70                  75                  80  


Cys Ala His Arg Thr Ser Leu Thr Asp Cys Gln Cys 
                85                  90          


<210>  24
<211>  84
<212>  PRT
<213>  Tetranychus urticae

<400>  24

Met Tyr Leu Lys Thr Tyr Phe Leu Leu Leu Val Ser Ile Phe Met Phe 
1               5                   10                  15      


Ser Leu Ile Thr Val Leu Leu Val Leu Pro Ser Leu Met Ala Ile Gln 
            20                  25                  30          


Thr Gly Cys Thr Asn Cys Ser Asn Cys Ile Asn Arg Thr Asn Cys His 
        35                  40                  45              


Gly Cys Glu Ser Cys Ser Asp Ser Thr Asn Cys Cys His Ser Arg Asn 
    50                  55                  60                  


Cys Ala Ser Cys Glu Tyr Leu Ser Thr Val Ser Thr Lys Leu Cys Arg 
65                  70                  75                  80  


Lys Phe Tyr His 
                


<210>  25
<211>  703
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Expression vector


<220>
<221>  MISC_FEATURE
<222>  (606)..(703)
<223>  AF protein

<400>  25

His Ala His Met His His His His His His Val Asn Asp Ala His Lys 
1               5                   10                  15      


Ser Glu Val Ala His Arg Phe Lys Asp Leu Gly Glu Glu Asn Phe Lys 
            20                  25                  30          


Ala Leu Val Leu Ile Ala Phe Ala Gln Tyr Leu Gln Gln Cys Pro Phe 
        35                  40                  45              


Glu Asp His Val Lys Leu Val Asn Glu Val Thr Glu Phe Ala Lys Thr 
    50                  55                  60                  


Cys Val Ala Asp Glu Ser Ala Glu Asn Cys Asp Lys Ser Leu His Thr 
65                  70                  75                  80  


Leu Phe Gly Asp Lys Leu Cys Thr Val Ala Thr Leu Arg Glu Thr Tyr 
                85                  90                  95      


Gly Glu Met Ala Asp Cys Cys Ala Lys Gln Glu Pro Glu Arg Asn Glu 
            100                 105                 110         


Cys Phe Leu Gln His Lys Asp Asp Asn Pro Asn Leu Pro Arg Leu Val 
        115                 120                 125             


Arg Pro Glu Val Asp Val Met Cys Thr Ala Phe His Asp Asn Glu Glu 
    130                 135                 140                 


Thr Phe Leu Lys Lys Tyr Leu Tyr Glu Ile Ala Arg Arg His Pro Tyr 
145                 150                 155                 160 


Phe Tyr Ala Pro Glu Leu Leu Phe Phe Ala Lys Arg Tyr Lys Ala Ala 
                165                 170                 175     


Phe Thr Glu Cys Cys Gln Ala Ala Asp Lys Ala Ala Cys Leu Leu Pro 
            180                 185                 190         


Lys Leu Asp Glu Leu Arg Asp Glu Gly Lys Ala Ser Ser Ala Lys Gln 
        195                 200                 205             


Arg Leu Lys Cys Ala Ser Leu Gln Lys Phe Gly Glu Arg Ala Phe Lys 
    210                 215                 220                 


Ala Trp Ala Val Ala Arg Leu Ser Gln Arg Phe Pro Lys Ala Glu Phe 
225                 230                 235                 240 


Ala Glu Val Ser Lys Leu Val Thr Asp Leu Thr Lys Val His Thr Glu 
                245                 250                 255     


Cys Cys His Gly Asp Leu Leu Glu Cys Ala Asp Asp Arg Ala Asp Leu 
            260                 265                 270         


Ala Lys Tyr Ile Cys Glu Asn Gln Asp Ser Ile Ser Ser Lys Leu Lys 
        275                 280                 285             


Glu Cys Cys Glu Lys Pro Leu Leu Glu Lys Ser His Cys Ile Ala Glu 
    290                 295                 300                 


Val Glu Asn Asp Glu Met Pro Ala Asp Leu Pro Ser Leu Ala Ala Asp 
305                 310                 315                 320 


Phe Val Glu Ser Lys Asp Val Cys Lys Asn Tyr Ala Glu Ala Lys Asp 
                325                 330                 335     


Val Phe Leu Gly Met Phe Leu Tyr Glu Tyr Ala Arg Arg His Pro Asp 
            340                 345                 350         


Tyr Ser Val Val Leu Leu Leu Arg Leu Ala Lys Thr Tyr Glu Thr Thr 
        355                 360                 365             


Leu Glu Lys Cys Cys Ala Ala Ala Asp Pro His Glu Cys Tyr Ala Lys 
    370                 375                 380                 


Val Phe Asp Glu Phe Lys Pro Leu Val Glu Glu Pro Gln Asn Leu Ile 
385                 390                 395                 400 


Lys Gln Asn Cys Glu Leu Phe Glu Gln Leu Gly Glu Tyr Lys Phe Gln 
                405                 410                 415     


Asn Ala Leu Leu Val Arg Tyr Thr Lys Lys Val Pro Gln Val Ser Thr 
            420                 425                 430         


Pro Thr Leu Val Glu Val Ser Arg Asn Leu Gly Lys Val Gly Ser Lys 
        435                 440                 445             


Cys Cys Lys His Pro Glu Ala Lys Arg Met Pro Cys Ala Glu Asp Tyr 
    450                 455                 460                 


Leu Ser Val Val Leu Asn Gln Leu Cys Val Leu His Glu Lys Thr Pro 
465                 470                 475                 480 


Val Ser Asp Arg Val Thr Lys Cys Cys Thr Glu Ser Leu Val Asn Arg 
                485                 490                 495     


Arg Pro Cys Phe Ser Ala Leu Glu Val Asp Glu Thr Tyr Val Pro Lys 
            500                 505                 510         


Glu Phe Asn Ala Glu Thr Phe Thr Phe His Ala Asp Ile Cys Thr Leu 
        515                 520                 525             


Ser Glu Lys Glu Arg Gln Ile Lys Lys Gln Thr Ala Leu Val Glu Leu 
    530                 535                 540                 


Val Lys His Lys Pro Lys Ala Thr Lys Glu Gln Leu Lys Ala Val Met 
545                 550                 555                 560 


Asp Asp Phe Ala Ala Phe Val Glu Lys Cys Cys Lys Ala Asp Asp Lys 
                565                 570                 575     


Glu Thr Cys Phe Ala Glu Glu Gly Lys Lys Leu Val Ala Ala Ser Gln 
            580                 585                 590         


Ala Ala Leu Gly Leu Gly Pro Gly Ser Asp Glu Val Asp Asp Gly Asn 
        595                 600                 605             


Thr Asn Cys Val Asn Cys Val Gly Cys Thr Asn Cys Ser Asn Cys Ile 
    610                 615                 620                 


Asn Cys Thr Asn Cys His Asp Cys Ala Asn Cys Thr Asp Ser Glu Asn 
625                 630                 635                 640 


Cys Ser Asn Ser Arg Asn Thr Ala Asn Ser Asn Gly Cys Gln Asn Cys 
                645                 650                 655     


Ala Asn Cys Thr Tyr Cys Val Ser Cys Ser Asp Cys Ala Asp Cys Ala 
            660                 665                 670         


Ser Ser Lys Gly Cys Ala Asn Cys Ala Asn Cys Thr Asn Cys Thr Asn 
        675                 680                 685             


Cys Arg Asn Cys Ala Asn Arg Ala Asn Leu Ser Asp Thr Gln Cys 
    690                 695                 700             


<210>  26
<211>  12
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Consensus repeat of insect antifreeze protein


<220>
<221>  misc_feature
<222>  (4)..(4)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (6)..(7)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (9)..(10)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (12)..(12)
<223>  Xaa can be any naturally occurring amino acid

<400>  26

Thr Cys Thr Xaa Ser Xaa Xaa Cys Xaa Xaa Ala Xaa 
1               5                   10          


<210>  27
<211>  12
<212>  PRT
<213>  Tetranychus urticae


<220>
<221>  misc_feature
<222>  (4)..(4)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (6)..(7)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (9)..(9)
<223>  Xaa can be any naturally occurring amino acid

<220>
<221>  misc_feature
<222>  (12)..(12)
<223>  Xaa can be any naturally occurring amino acid

<400>  27

Asn Cys Thr Xaa Cys Xaa Xaa Cys Xaa Asn Cys Xaa 
1               5                   10          


