Trypsin

one member of the serine protease family

plays important roles in animal protein digestion. It is synthesized and secreted as proenzyme trypsinogen

and removed the N-terminal activation peptide by enterokinase converting to active form in the intestine. The active trypsin can specifically cleavage at the peptide bond on the carboxyl side of basic L-amino acids such as arginine or lysine residuse. For studying the protein structure and physiological funtions of trypsinogen in fish

we successfully got three trypsinogen cDNAs (zftry1a、zftry1b and zftry2) from zebrafish by RT-PCR and RACE. The results showed zftry1a and zftry1b consisted of 242 amino acids which contained a signal peptide(SP) of 15 amino acids and an activation peptide(AP) of five amino acids

LDDDK. zftry2 consisted of 247 amino acids which contained a SP of 15 amino acids and an AP of nine amino acids

APLGDDDDK. The alignment based on the amino acid sequences revealed that they had the conversed structural characteristics

such as the catalytic triad (His-57

Asp-102

and Ser-195)

12 cysteines forming 6 disulfide bonds

Asp-189 at the bottom of the substrate-binding pocket and Gly-216 and Gly-226 lining the sides of the binding pocket

and so on. The results from isoelectric and phylogenetic analyses suggested that zftry1a and zftry1b were group with teleost anionic trypsinogen group I

while zftry2 was phylogenetically related to teleost cationic group II. Tissue expression pattern was similar to each other

and all trypsinogens were mainly expressed in the intestine. These results from zebrafish trypsinogens can provide the foundations for further study of its expression profiles

the molecular characteristics and evolution of fish trypsinogen.