Cloning and characterization of a β-1,4-mannanase 5C possessing a family 27 carbohydrate-binding module from a marine bacterium, Vibrio sp. strain MA-138

Abstract

The β-1,4-mannanase 5C gene (man5C) of Vibrio sp. strain MA-138 was cloned and expressed in Escherichia coli. The man5C gene consisted of 2,010 bp nucleotides encoding a protein of 669 amino acids with a predicted molecular weight of 76,309. β-1,4-Mannanase (Man5C) is a modular enzyme composed of a catalytic module belonging to glycoside hydrolase family 5, a linker region, and a putative carbohydrate-binding module (CBM) belonging to family 27. Recombinant Man5C exhibited maximal activity at 50°C at pH 7.0, and it had a Km of 0.6 mg ml-1 and a Vmax of 556.2 μmol min-1 μmol-1 for glucomannan. Binding studies revealed that the C-terminal putative CBM27 had the ability to bind soluble β-mannans and contributed to increasing the rate of depolymerization by binding to the polymeric substrate. Man5C of Vibrio sp. MA-138 is the first non-extremophile enzyme to be identified as a β-mannanase possessing CBM27.