[Isolation and properties of carboxypeptidase from the Kamchatka crab Paralithodes camtshatica].
Homogeneous carboxypeptidase PC from a hematopancreas of kamchatka crab Paralithodes camtshatica was obtained by means of an affinity chromatography on sorbents containing arginine, protamine hydrolysate, and phenylalanine as ligands with an yield 23% and purification degree 37.4. The isolated enzyme has a molecular mass 34 kDa, as evidenced by an SDS-PAGE; pI 3.1; an optimum pH 6.5, as estimated for hydrolysis of Dnp-Ala-Ala-Arg; pH-stability range 5-8 in the presence of Ca2+; a temperature optimum 55 degrees C; and Km 0.4 mM. The carboxypeptidase is activated by Co2+ and Ca2+ ions and is inhibited by EDTA and o-phenanthroline, and therefore, it is a metallocarboxypeptidase. The enzyme can effectively split off C-terminal residues Phe and Tyr, as well as Arg and Lys. Residues Pro, Glu, and Asp cannot be split off, and they stop the cleaving of a preceding bond. Thus, the carboxypeptidase PC of kamchatka crab has a mixed substrate specificity, which is characteristic of carboxypeptidase from crawfish and of microbial carboxypeptidases T and SG. The new carboxypeptidase has an amino acid composition Asp41Thr24Ser22Glu32Pro15Gly32Ala291/2Cys5Val19Met8Ile14Leu20Ty r18Phe8Lys7His4 Arg8Trp4. The N-terminal sequence of the enzyme demonstrate a 40% homology with the N-terminal sequence of carboxypeptidase from crawfish.
Rudenskaia
G N
GN
Kupenko
O G
OG
Isaev
V A
VA
Stepanov
V M
VM
Dunaevskiĭ
Ia E
IaE
D000595:Amino Acid Sequence; D000818:Animals; D003386:Brachyura / Q000201:enzymology*; D002268:Carboxypeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000302:isolation & purification*; D002846:Chromatography, Affinity; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity
D002268:Carboxypeptidases; C094140:carboxypeptidase PC
Vydelenie i svoĭstva karboksipeptidazy kamchatskogo kraba Paralithodes camtshatica.