Possible involvement of an FKBP family member protein from a psychrotrophic bacterium Shewanella sp. SIB1 in cold-adaptation

Eur J Biochem. 2004 Apr;271(7):1372-81. doi: 10.1111/j.1432-1033.2004.04049.x.

Abstract

A psychrotrophic bacterium Shewanella sp. strain SIB1 was grown at 4 and 20 degrees C, and total soluble proteins extracted from the cells were analyzed by two-dimensional polyacrylamide gel electrophoresis. Comparison of these patterns showed that the cellular content of a protein with a molecular mass of 28 kDa and an isoelectric point of four greatly increased at 4 degrees C compared to that at 20 degrees C. Determination of the N-terminal amino acid sequence, followed by the cloning and sequencing of the gene encoding this protein, revealed that this protein is a member of the FKBP family of proteins with an amino acid sequence identity of 56% to Escherichia coli FKBP22. This protein was overproduced in E. coli in a His-tagged form, purified, and analyzed for peptidyl-prolyl cis-trans isomerase activity. When this activity was determined by the protease coupling assay using N-succinyl-Ala-Leu-Pro-Phe-p-nitroanilide as a substrate at various temperatures, the protein exhibited the highest activity at 10 degrees C with a k(cat)/K(m) value of 0.87 micro m(-1) x s(-1). When the peptidyl-prolyl cis-trans isomerase activity was determined by the RNase T(1) refolding assay at 10 and 20 degrees C, the protein exhibited higher activity at 10 degrees C with a k(cat)/K(m) value of 0.50 micro m(-1) x s(-1). These k(cat)/K(m) values are lower but comparable to those of E. coli FKBP22. We propose that a FKBP family protein is involved in cold-adaptation of psychrotrophic bacteria.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Catalysis
  • Cloning, Molecular
  • Cold Temperature*
  • DNA / chemistry
  • Electrophoresis, Gel, Two-Dimensional
  • Electrophoresis, Polyacrylamide Gel
  • Escherichia coli / metabolism
  • Hydrogen-Ion Concentration
  • Kinetics
  • Models, Genetic
  • Molecular Sequence Data
  • Molecular Weight
  • Peptidylprolyl Isomerase / chemistry
  • Plasmids / metabolism
  • Protein Binding
  • Protein Denaturation
  • Protein Folding
  • Protein Structure, Tertiary
  • Recombinant Proteins / chemistry
  • Ribonuclease T1 / chemistry
  • Sequence Homology, Amino Acid
  • Shewanella / metabolism*
  • Tacrolimus Binding Proteins / chemistry*
  • Temperature
  • Time Factors

Substances

  • Recombinant Proteins
  • DNA
  • Ribonuclease T1
  • FKBP22
  • Tacrolimus Binding Proteins
  • Peptidylprolyl Isomerase

Associated data

  • GENBANK/AB116100