Molecular Identification of Family 38 α-Mannosidase of Bacillus sp. Strain GL1, Responsible for Complete Depolymerization of Xanthan

Abstract

ABSTRACT When cells of Bacillus sp. strain GL1 were grown in a medium containing xanthan as a carbon source, α-mannosidase exhibiting activity toward p -nitrophenyl-α- d -mannopyranoside ( p NP-α- d -Man) was produced intracellularly. The 350-kDa α-mannosidase purified from a cell extract of the bacterium was a trimer comprising three identical subunits, each with a molecular mass of 110 kDa. The enzyme hydrolyzed p NP-α- d -Man ( K m = 0.49 mM) and d -mannosyl-(α-1,3)- d -glucose most efficiently at pH 7.5 to 9.0, indicating that the enzyme catalyzes the last step of the xanthan depolymerization pathway of Bacillus sp. strain GL1. The gene for α-mannosidase cloned most by using N-terminal amino acid sequence information contained an open reading frame (3,144 bp) capable of coding for a polypeptide with a molecular weight of 119,239. The deduced amino acid sequence showed homology with the amino acid sequences of α-mannosidases belonging to glycoside hydrolase family 38.