Molecular and enzymatic properties of a cathepsin L-like proteinase with distinct substrate specificity from northern shrimp (Pandalus borealis)

被引:14
作者
Aoki, H
Ahsan, MN
Watabe, S
机构
[1] Univ Tokyo, Grad Sch Agr & Life Sci, Lab Aquat Mol Biol & Biotechnol, Bunkyo Ku, Tokyo 1138657, Japan
[2] Nichirei Corp, Ctr Res & Dev, Mihama Ku, Chiba 2618545, Japan
关键词
cathepsin L; cysteine proteinase; S2; subsite; Northern shrimp; Pandalus borealis;
D O I
10.1007/s00360-003-0389-x
中图分类号
Q4 [生理学];
学科分类号
071003 ;
摘要
We purified a cathepsin L-like proteinase to homogeneity from the hepatopancreas of northern shrimp Pandalus borealis by several chromatographic procedures. The purified proteinase showed the highest specificity for leucine residue at P2, a specificity pattern similar to cathepsins S and K whereas proline and arginine residues were not suitable as P2 substrates. However, unlike these proteinases, it accepted valine almost equally to the phenylalanine residue at P2. The shrimp cathepsin was strongly inhibited by E-64, leupeptin and antipain, while benzyloxycarbonyl-Phe-Tyr(t-Bu)-CHN2, a specific inhibitor of cathepsin L, remained largely ineffective. Next, we determined the primary structure of the shrimp enzyme by molecular cloning and investigated the residues constituting the S2 subsite, which is possibly involved in its unusual substrate specificity. The deduced amino acid sequence of the shrimp proteinase shared the highest identity of 65% with a cathepsin L-like proteinase from lobster, but its identity to the well-characterized mammalian cathepsins S, L, and K fell within narrower ranges of 52-55%. However, the shrimp proteinase differed from these cathepsins in some key residues including, for example, the unique occurrence of cysteine and glutamine residues at the structurally important S2 subsite. Interestingly, transcripts of this proteinase were exclusively detected in the shrimp gut coinciding with its broad pH activity and stability profiles, which is also unusual as a cysteine proteinase. These results suggest that the shrimp enzyme is homologous to mammalian cathepsins S, L, and K, but is distinct from each of these proteinases in both enzymatic and structural properties.
引用
收藏
页码:59 / 69
页数:11
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