Stability of protease in organic solvent: Structural identification by solid-state NMR of lyophilized papain before and after 1-propanol treatment and the corresponding enzymatic activities

Teruhiko Matsubara, Risa Fujita, Shigeru Sugiyama, Katsuhiro Kawashiro

Research output: Contribution to journalArticle

2 Citations (Scopus)

Abstract

Lyophilized enzyme powder is often used in organic solvents. However, the enzymatic activity decreases during the reaction process. In the present study, the relation between structural stability and enzymatic activity in an organic solvent was investigated. 13C cross-polarization magic angle spinning NMR spectroscopy was used to determine the secondary structure of lyophilized papain in the solid-state. Deconvolution of the peaks of the backbone carbonyl carbons suggested that the proportion of β-sheet conformation increased after lyophilization from a phosphate buffer solution. The esterification of N-benzyloxycarbonyl phenylalanylalanine amide was attempted using the lyophilized papain as a catalyst in anhydrous 1-propanol. The yield of ester was 46.1% after 48 h at 50°C, but this reaction slowed remarkably after 48 h. When the lyophilized papain was suspended in anhydrous 1-propanol for 7 days without the substrate, the proportion of β-sheet conformation was further increased and the suspended papain had no activity. These results suggest that the increase in β-sheet conformation caused inactivation of papain. The increase in β-sheet conformation caused by both lyophilization and suspension in propanol was found, which was related to a decrease in enzymatic activity.

Original languageEnglish
Pages (from-to)928-933
Number of pages6
JournalBiotechnology and bioengineering
Volume93
Issue number5
DOIs
Publication statusPublished - 2006 Apr 5

Keywords

  • CPMAS solid-state NMR
  • Lyophilization
  • Organic solvent
  • Papain
  • Structural stability

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Applied Microbiology and Biotechnology

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