Biochemistry (Moscow)

, Volume 71, Issue 5, pp 563–570

Psychrophilic trypsin-type protease from Serratia proteamaculans

  • A. G. Mikhailova
  • V. V. Likhareva
  • R. F. Khairullin
  • N. L. Lubenets
  • L. D. Rumsh
  • I. V. Demidyuk
  • S. V. Kostrov
Article

Abstract

A preparative method for purification of a novel protease from the psychrotolerant Gram-negative microorganism Serratia proteamaculans (PSP) was developed using affinity chromatography on BPTI-Sepharose. It yielded electrophoretically homogeneous PSP preparation of 60 kD. The PSP properties (temperature and pH stability, high catalytic efficiency) indicate that this enzyme can be defined as a psychrophilic protease. Inhibitory analysis together with substrate specificity indicates that the studied PSP exhibits properties of serine trypsin-like and Zn-dependent protease.

Key words

psychrophilic enzymes trypsin Serratia proteamaculans affinity chromatography substrate-inhibitor analysis 

Abbreviations

ATEE

Nα-acetyl-L-tyrosine ethyl ester

BAPNA

Nα-benzoyl-DL-arginine p-nitroanilide

BPTI

bovine pancreatic trypsin inhibitor

DFP

diisopropylfluorophosphate

DMSO

dimethylsulfoxide

DTDP

4,4′-dithiodipyridine

PMSF

phenylmethylsulfonyl fluoride

PSP

Serratia proteamaculans protease

STI

soybean trypsin inhibitor

TLCK

Nα-p-tosyl-L-lysyl-chloromethyl-ketone

Z-Lys-S-Bzl

Nα-benzyloxycarbonyl-L-lysine thiobenzyl ester

buffer A

0.1 M Tris-HCl, pH 8.0, 50 mM CaCl2

buffer B

0.1 M Tris-HCl, pH 8.0

buffer C

0.1 M Tris-HCl, pH 9.0, 50 mM CaCl2, 1 mM MgCl2

buffer D

10 mM Hepes-KOH, pH 7.5, 1 mM MgCl2

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Copyright information

© Pleiades Publishing, Inc. 2006

Authors and Affiliations

  • A. G. Mikhailova
    • 1
  • V. V. Likhareva
    • 1
  • R. F. Khairullin
    • 1
  • N. L. Lubenets
    • 1
  • L. D. Rumsh
    • 1
  • I. V. Demidyuk
    • 2
  • S. V. Kostrov
    • 2
  1. 1.Shemyakin and Ovchinnikov Institute of Bioorganic ChemistryRussian Academy of SciencesMoscowRussia
  2. 2.Institute of Molecular GeneticsRussian Academy of SciencesMoscowRussia

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