Characterization of a native cellulase activity from an anaerobic thermophilic hydrogen-producing bacterium Thermosipho sp. strain 3

Abstract

A bacterial strain, designated as strain 3 and identified as a member of the Thermosipho species on the basis of its phenotypic and genotypic characteristics, was isolated from a deep sea hydrothermal vent. Sequence analysis of the 16S rRNA gene revealed that its closest neighbor was Thermosipho africanus (99.5 %). This isolate Thermosipho sp. strain 3 (DSM 27729), a thermophilic, anaerobic, fermentative hydrogen-producing bacterium, produced a thermostable endocellulase that hydrolyzes carboxymethylcellulose (CMC) and β-glucan. The cellulase was purified and its activity characterized. The estimated molecular weight of the protein was about 40 kDa as determined by gel-filtration chromatography, SDS-PAGE and zymogram analyses. The optimal cellulase activity was at pH 5.5 and at a temperature of 80 °C. The enzyme was thermostable with about 50 % residual activity after 48 h and 4 h at 60 °C and 70 °C, respectively. Interestingly, endocellulase activity was increased about 2-fold by 5 mM MnCl2. MALDI-TOF PMF and the N-terminal amino acid sequence analyses of the purified enzyme revealed the extensive homology of the protein with a glycoside hydrolase family protein from Thermosipho africanus (NCBI protein accession number: 419759359; UniProt: K2PFP0).