The propeptide of subtilisin BPN′ as a temporary inhibitor and effect of an amino acid replacement on its inhibitory activity

FEBS Letters

Miura, Kin-ichiro¹  Kojima, Shuichi¹  Minagawa, Tetsuya¹ 

[1] Institute for Biomolecular Science, Gakushuin University, Mejiro, Tokyo 171, Japan

关键词: Amino acid replacement;    Propeptide;    protease inhibitor;    Subtilisin;    Temporary inhibition;   

DOI  :  10.1016/S0014-5793(97)00678-9

学科分类：生物化学/生物物理

来源: John Wiley & Sons Ltd.

【 摘 要 】

The propeptide of subtilisin-family proteases is known to exhibit inhibitory activity toward a cognate protease in addition to its function as an intramolecular chaperone. For detailed investigation of its inhibitory properties, the propeptide of subtilisin BPN′ was produced in Escherichia coli. Inhibitory activity measurements and electrophoresis showed that the propeptide was a temporary inhibitor, which was initially potent but was gradually degraded by subtilisin BPN′ through specific intermediates. The main cleavage site was identified as Glu⁵³–Lys⁵⁴, with minor sites at Thr¹⁷–Met¹⁸ and Met²¹–Ser²², which were located in turn regions of the propeptide in the complex with subtilisin BPN′. Since the isolated propeptide has been shown not to form a tertiary structure, these results indicate that main digestions proceed through proteolytic attack of subtilisin toward the accessible sites of the propeptide in the complex with subtilisin. Therefore, replacement of Glu53 at the main cleavage site by Asp, which is a less favorable amino acid than Glu for subtilisin, makes the propeptide a more resistant temporary inhibitor.

【 授权许可】

Unknown