Functional analysis of the intramolecular chaperone. Mutational hot spots in the subtilisin pro-peptide and a secondsite suppressor mutation within the subtilisin molecule

Tatsuhiko Kobayashi, Masayori Inouye

Research output: Contribution to journalArticlepeer-review

40 Scopus citations

Abstract

The N-terminal pro-peptide of 77 amino acid residues is essential for the folding of subtilisin, an alkaline serine protease from Bacillus subtilis. The synthetic pro-peptide has been shown to be capable of guiding the proper folding of denatured subtilisin to enzymatically active enzyme. Thus the pro-peptide serves as an intramolecular chaperone, which is removed by an autoprocessing reaction after the completion of the folding. With use of localized polymerase chain reaction random mutagenesis a total of 25 amino acid substitution mutations that affected subtilisin activities were isolated. These mutations occurred in a high frequency at the hydrophobic regions of the pro-peptide. For one of the mutations, M(-60)T, a second-site suppressor mutation, S(188)L, was isolated within the mature region. These results suggest that the pro-peptide consists of a few functional regions which interact with specific regions of the mature region of subtilisin during the folding process.

Original languageEnglish (US)
Pages (from-to)931-933
Number of pages3
JournalJournal of molecular biology
Volume226
Issue number4
DOIs
StatePublished - May 20 1992

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Molecular Biology

Keywords

  • chaperone
  • pro-peptide
  • protein folding
  • subtilisin

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