                         SEQUENCE LISTING

<110>  UNIVERSITE LAVAL
       SALESSE, Christian
       CANTIN, Line
 
<120>  RECOVERIN AS A FUSION PROTEIN TAG TO IMPROVE EXPRESSION, 
       SOLUBILITY AND PURIFICATION OF PROTEINS

<130>  000819-0318

<150>  62/194,348
<151>  2015-07-20

<160>  4     

<170>  PatentIn version 3.5

<210>  1
<211>  609
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Nucleic acid sequence - full length native Recoverin

<400>  1
atggggaaca gcaagagtgg ggccctgtcc aaggagatcc tggaggagct gcagctgaac       60

accaagttca cggaggagga gctgagctcc tggtaccagt ccttcctgaa ggagtgcccc      120

agtggccgga tcacccggca ggagttccag accatctact ccaagttctt ccccgaggcc      180

gaccccaagg cctatgccca gcacgtgttc cgaagctttg atgccaacag cgatggcacc      240

ttggacttca aggagtatgt catcgcccta cacatgacca gcgcgggcaa gaccaaccag      300

aagctggagt gggccttctc cctctatgat gtggatggca atgggaccat cagcaagaac      360

gaggtgctgg agattgtcac ggctatcttc aaaatgatca gccctgagga cacaaagcat      420

ctcccagaag acgagaacac tccggaaaag cgagcagaga agatctgggg attctttggc      480

aagaaggatg atgataaact tacagagaaa gaattcatcg aagggaccct ggccaataag      540

gaaattctgc gactgattca attcgagcct caaaaagtga aggagaaact gaaggaaaag      600

aaactctga                                                              609


<210>  2
<211>  201
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Amino acid sequence - full length native Recoverin (start Met is 
       removed)

<400>  2

Gly Asn Ser Lys Ser Gly Ala Leu Ser Lys Glu Ile Leu Glu Glu Leu 
1               5                   10                  15      


Gln Leu Asn Thr Lys Phe Thr Glu Glu Glu Leu Ser Ser Trp Tyr Gln 
            20                  25                  30          


Ser Phe Leu Lys Glu Cys Pro Ser Gly Arg Ile Thr Arg Gln Glu Phe 
        35                  40                  45              


Gln Thr Ile Tyr Ser Lys Phe Phe Pro Glu Ala Asp Pro Lys Ala Tyr 
    50                  55                  60                  


Ala Gln His Val Phe Arg Ser Phe Asp Ala Asn Ser Asp Gly Thr Leu 
65                  70                  75                  80  


Asp Phe Lys Glu Tyr Val Ile Ala Leu His Met Thr Ser Ala Gly Lys 
                85                  90                  95      


Thr Asn Gln Lys Leu Glu Trp Ala Phe Ser Leu Tyr Asp Val Asp Gly 
            100                 105                 110         


Asn Gly Thr Ile Ser Lys Asn Glu Val Leu Glu Ile Val Thr Ala Ile 
        115                 120                 125             


Phe Lys Met Ile Ser Pro Glu Asp Thr Lys His Leu Pro Glu Asp Glu 
    130                 135                 140                 


Asn Thr Pro Glu Lys Arg Ala Glu Lys Ile Trp Gly Phe Phe Gly Lys 
145                 150                 155                 160 


Lys Asp Asp Asp Lys Leu Thr Glu Lys Glu Phe Ile Glu Gly Thr Leu 
                165                 170                 175     


Ala Asn Lys Glu Ile Leu Arg Leu Ile Gln Phe Glu Pro Gln Lys Val 
            180                 185                 190         


Lys Glu Lys Leu Lys Glu Lys Lys Leu 
        195                 200     


<210>  3
<211>  642
<212>  DNA
<213>  Artificial Sequence

<220>
<223>  Nucleic acid sequence - mutated Recoverin (+ NdeI and BamHI 
       restriction sites, + silent mutation, without myristoyl group)


<220>
<221>  misc_feature
<222>  (1)..(6)
<223>  Nde1 site (CATATG)

<220>
<221>  misc_feature
<222>  (333)..(333)
<223>  T to C: silent mutation

<220>
<221>  misc_feature
<222>  (610)..(636)
<223>  5 glycines (GGC or GGT) at the end  + thrombin cleavage site 
       [Leu-Val-Pro-Arg-Gly-Ser : L (CTG), V (GTT), P (CCG), R (CGT), G 
       (GGA), S (TCC)]

<220>
<221>  misc_feature
<222>  (637)..(642)
<223>  Stop codon removed and replaced by BamH1 site (GGATTC) that 
       corresponds also to glycine and serine from thrombin recognition 
       site

<400>  3
catatgggga acagcaagag tggggccctg tccaaggaga tcctggagga gctgcagctg       60

aacaccaagt tcacggagga ggagctgagc tcctggtacc agtccttcct gaaggagtgc      120

cccagtggcc ggatcacccg gcaggagttc cagaccatct actccaagtt cttccccgag      180

gccgacccca aggcctatgc ccagcacgtg ttccgaagct ttgatgccaa cagcgatggc      240

accttggact tcaaggagta tgtcatcgcc ctacacatga ccagcgcggg caagaccaac      300

cagaagctgg agtgggcctt ctccctctat gacgtggatg gcaatgggac catcagcaag      360

aacgaggtgc tggagattgt cacggctatc ttcaaaatga tcagccctga ggacacaaag      420

catctcccag aagacgagaa cactccggaa aagcgagcag agaagatctg gggattcttt      480

ggcaagaagg atgatgataa acttacagag aaagagttca tcgaagggac cctggccaat      540

aaggaaattc tgcgactgat tcaattcgag cctcaaaaag tgaaggagaa actgaaggaa      600

aagaaactcg gcggtggtgg cggcctggtt ccgcgtggat cc                         642


<210>  4
<211>  212
<212>  PRT
<213>  Artificial Sequence

<220>
<223>  Amino acid sequence - mutated Recoverin (+ NdeI and BamHI 
       restriction sites, - EcoR1 silent mutation)


<220>
<221>  MISC_FEATURE
<222>  (202)..(212)
<223>  Thrombin cleavage site

<400>  4

Gly Asn Ser Lys Ser Gly Ala Leu Ser Lys Glu Ile Leu Glu Glu Leu 
1               5                   10                  15      


Gln Leu Asn Thr Lys Phe Thr Glu Glu Glu Leu Ser Ser Trp Tyr Gln 
            20                  25                  30          


Ser Phe Leu Lys Glu Cys Pro Ser Gly Arg Ile Thr Arg Gln Glu Phe 
        35                  40                  45              


Gln Thr Ile Tyr Ser Lys Phe Phe Pro Glu Ala Asp Pro Lys Ala Tyr 
    50                  55                  60                  


Ala Gln His Val Phe Arg Ser Phe Asp Ala Asn Ser Asp Gly Thr Leu 
65                  70                  75                  80  


Asp Phe Lys Glu Tyr Val Ile Ala Leu His Met Thr Ser Ala Gly Lys 
                85                  90                  95      


Thr Asn Gln Lys Leu Glu Trp Ala Phe Ser Leu Tyr Asp Val Asp Gly 
            100                 105                 110         


Asn Gly Thr Ile Ser Lys Asn Glu Val Leu Glu Ile Val Thr Ala Ile 
        115                 120                 125             


Phe Lys Met Ile Ser Pro Glu Asp Thr Lys His Leu Pro Glu Asp Glu 
    130                 135                 140                 


Asn Thr Pro Glu Lys Arg Ala Glu Lys Ile Trp Gly Phe Phe Gly Lys 
145                 150                 155                 160 


Lys Asp Asp Asp Lys Leu Thr Glu Lys Glu Phe Ile Glu Gly Thr Leu 
                165                 170                 175     


Ala Asn Lys Glu Ile Leu Arg Leu Ile Gln Phe Glu Pro Gln Lys Val 
            180                 185                 190         


Lys Glu Lys Leu Lys Glu Lys Lys Leu Gly Gly Gly Gly Gly Leu Val 
        195                 200                 205             


Pro Arg Gly Ser 
    210         


