SEQUENCING, CLONING AND EXPRESSION OF A BETA-1,4-MANNANASE GENE, MANA, FROM THE EXTREMELY THERMOPHILIC ANAEROBIC BACTERIUM, CALDICELLULOSKIRUPTOR RT8B.4

A gene encoding a beta-mannanase (manA) has been cloned from an obliga tely anaerobic extreme thermophile, Caldicellulosiruptor strain Rt8B.4 , which is most closely related to Caldicellulosiruptor saccharolyticu s (formerly Caldocellum saccharolyticum). The gene codes for a multido main enzyme with a C-terminal beta-mannanase domain which was amplifie d by the polymerase chain reaction and cloned into a temperature-induc ible expression vector in Escherichia coil. Sequence comparisons have shown that the Man domain of Rt8B.4 ManA is related to a thermophilic Dictyoglomus mannanase and a mesophilic mannanase from a Bacillus spec ies. It appears to be unrelated to the beta-mannanase domain of C. sac charolyticus, implying acquisition of the genes from unrelated sources by the two bacteria.