Intermolecular autocatalytic activation of serine protease zymogen factor C through an active transition state responding to lipopolysaccharide

Toshio Shibata, Yuki Kobayashi, Yuto Ikeda, Shun ichiro Kawabata

Research output: Contribution to journalArticle

4 Citations (Scopus)

Abstract

Horseshoe crab hemolymph coagulation is believed to be triggered by the autocatalytic activation of serine protease zymogen factor C to the active form, -factor C, belonging to the trypsin family, through an active transition state of factor C responding to bacterial lipopolysaccharide (LPS), designated factor C*. However, the existence of factor C* is only speculative, and its proteolytic activity has not been validated. In addition, it remains unclear whether the proteolytic cleavage of the Phe737–Ile738 bond (Phe737 site) of factor C required for the conversion to -factor C occurs intramolecularly or intermolecularly between the factor C molecules. Here we show that the Phe737 site of a catalytic Ser-deficient mutant of factor C is LPS-dependently hydrolyzed by a Phe737 site– uncleavable mutant, clearly indicating the existence of the active transition state of factor C without cleavage of the Phe737 site. Moreover, we found the following facts using several mutants of factor C: the autocatalytic cleavage of factor C occurs intermolecularly between factor C* molecules on the LPS surface; factor C* does not exhibit intrinsic chymotryptic activity against the Phe737 site, but it may recognize a three-dimensional structure around the cleavage site; and LPS is required not only to complete the substrate-binding site and oxyanion hole of factor C* by interacting with the N-terminal region but also to allow the Phe737 site to be cleaved by inducing a conformational change around the Phe737 site or by acting as a scaffold to induce specific protein–protein interactions between factor C* molecules.

Original languageEnglish
Pages (from-to)11589-11599
Number of pages11
JournalJournal of Biological Chemistry
Volume293
Issue number29
DOIs
Publication statusPublished - Jul 20 2018

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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