Thermotoga neapolitana bgIB gene, upstream of lamA, encodes a highly thermostable β-glucosidase that is a laminaribiase

Abstract

The gene for thermostable 1,3-β-glucosidase BgIB was cloned from the chromosome of Thermotoga neapolitana and its primary sequence was determined. The purified recombinant β-glucosidase B had a monomer molecular mass of 81 kDa in accordance with the amino acid sequence predicted from the nucleotide sequence of clone pTT51. It was a member of glycosylhydrolase family 3 and belonged to enzyme class EC 3.2.1.21. β-Glucosidase B had a specific activity of 255 U mg-1on 4-nitrophenyl(PNP)-β-glucoside at the optima of pH (5.5) and temperature (90 °C), and K m values of 0.1, 10 and 50 mM for PNP-β-glucoside, laminaribiose and cellobiose, respectively. The gene bgIB was located immediately upstream of the laminarinase gene IamA. Both genes were transcribed from the same DNA strand and were not separated by a palindromic transcription terminator. The two purified enzymes 1,3-β-glucosidase BgIB (laminaribiase) and 1,3-β-glucanase LamA (laminarinase) were together capable of completely degrading laminarin to glucose.