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

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 45kDa 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 K(m) measured for the CEDAase was 55.6μM, with an apparent V(max) of 0.083μmol/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.