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The Protein Journal

, Volume 31, Issue 1, pp 51–58 | Cite as

Purification and Characterization of an Endo-d-arabinase Produced by Cellulomonas

  • Ming-Zhong Sun
  • Xiao-Ying Zhang
  • Yi Xin
Article

Abstract

An extracellular endo-d-arabinase enzyme produced by the bacterial strain of Cellulomonas was purified 77.1-fold with 0.20% recovery for protein by DEAE Sepharose anion exchange, Sephacryl S-300 gel filtration and blue Sepharose affinity chromatography, and designated as CEDAase. The apparent molecular mass of CEDAase was 45 kDa determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. CEDAase is an endoenzyme for arabinogalactan with the main and specific product of hexa-arabinofuranoside. It reacts optimally with its substrate, arabinogalactan, at approximately pH 8.0 and at 40 °C. CEDAase shows stability in the pH range of 6.0–9.0 and at the temperature below 50 °C. The Km measured for the CEDAase was 55.6 μM, with an apparent Vmax of 0.083 μmol/min. To our knowledge, for the first time, the current work obtains an extracellular Cellulomonas endo-d-arabinase enzyme that might be potentially served as a tool enzyme for hydrolyzing specific cell wall such as Mycobacterium cell. It is purified as an important potential initial material basis for mass spectrometric sequencing and chemical gene synthesis. It may make it possible to clone and express this valuable endo-d-arabinase and make it available to the mycobacteria scientific community.

Keywords

Endo-arabinase enzyme Cellulomonas Purification Characterization 

Abbreviations

AG

Arabinogalactan

CEDAase

Cellulomonas endo-d-arabinase

DEAE Sepharose

Diethylaminoethyl Sepharose

SDS-PAGE

Sodium dodecyl sulfate-polyacrylamide gel electrophoresis

TB

Tuberculosis

MDR

Multiple drugs resistant (or Multiple drugs resistance)

PBS

Phosphate buffer solution

HAC

Acetic acid

TLC

Thin layer chromatography

HPLC

High performance liquid chromatography

PAD

Pulsed amperometric detector

Notes

Acknowledgments

This work was supported by a grant from National Natural Science Foundation of China (306570416).

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Copyright information

© Springer Science+Business Media, LLC 2011

Authors and Affiliations

  1. 1.Department of BiotechnologyDalian Medical UniversityDalianChina

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