Characterization of Mannose Isomerase from a Cellulolytic Actinobacteria Thermobifida fusca MBL10003

抄録

D-mannose isomerase was cloned and characterized from a newly isolated actinobacteria strain, Thermobifida fusca MBL10003. The structural gene (manI) is predicted to encode a polypeptide of 407 amino acids with an estimated molecular mass of 43,900. Although the identity of the deduced amino acid sequence is not so high (45.7% to Salmonella enterica, and 38.5% to Agrobacterium radiobacter), the catalytic center appears has an essential structure that is conserved in the characterized homologs. It was a dimeric enzyme composed of two active monomer units. The optimal temperature and pH were 60°C and 8.0, respectively. The enzyme was stable up to around 60°C, and between pH 4 and 11. It showed activity on D-mannose and also on D-lyxose, but not on N-acetyl D-glucosamine, suggesting that it is functionally different from N-acyl D-glucosamine 2-epimerase despite the sequence similarity. Although K_m value for D-mannose, 115 mM was similar to other mannose isomerase, k_cat, hence k_cat/K_m was much higher than those. The enzyme was significantly inhibited by such divalent metal ions as Cu²⁺, Cd²⁺ or Ca²⁺, but it was not a metal-required enzyme for activity emergence.