Purification and characterization of hatching enzyme from shrimp Penaeus chinensis.
By using Penaeus chorion as a specific substrate, the hatching enzyme (HE) from Penaeus chinensis was purified by gel-filtration and ion-exchange chromatography, and characterized in terms of its molecular weight and enzymatic properties in this study. It was found that the molecular weight of Penaeus HE is about 43.0 kDa in SDS-PAGE. The Penaeus HE had obvious choriolytic activity, which was optimal at pH 6.0 and temperature of 40 degrees C, respectively. The Km value of the HE for casein was 7.47 mg ml(-1). The HE activity was almost completely inhibited by SBTI, p-APMSF, bestatin, and NEM, greatly inhibited by ovomucoid, TLCK, IAM, chymostatin, and PMSF, and slightly inhibited by pepstatin A, TPCK, LBTI, and leupeptin. These results indicate that the HE is most probably a trypsin-type serine protease. Besides of these, the HE was extremely sensitive to EDTA, Zn2+, Ca2+, Mg2+, and Cu2+. Combined with the results that the EDTA-pretreated HE activity could be perfectly recovered by Zn2+, it is indicated that shrimp HE is most probably a kind of Zn-metalloprotease.
Li
Bing-Jun
BJ
Department of Marine Biology, College of Marine Life Sciences, Ocean University of China, Qingdao 266003, China.
Fan
Ting-Jun
TJ
Yang
Ling-Ling
LL
Cong
Ri-Shan
RS
Li
Ling
L
Sun
Wen-Jie
WJ
Lu
Cui-Xian
CX
Shi
Zhen-Ping
ZP
D000818:Animals; D002364:Caseins / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D002823:Chorion / Q000737:chemistry; D004492:Edetic Acid / Q000494:pharmacology; D004625:Embryo, Nonmammalian; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007477:Ions / Q000494:pharmacology; D007700:Kinetics; D008670:Metals / Q000494:pharmacology; D008970:Molecular Weight; D033561:Penaeidae / Q000196:embryology* / Q000201:enzymology*; D012697:Serine Endopeptidases / Q000032:analysis / Q000737:chemistry* / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity; D013696:Temperature
D002364:Caseins; D002614:Chelating Agents; D004791:Enzyme Inhibitors; D007477:Ions; D008670:Metals; D004492:Edetic Acid; D012697:Serine Endopeptidases