Screening and evolution of a novel protist xylose isomerase from the termite Reticulitermes speratus for efficient xylose fermentation in Saccharomyces cerevisiae

The yeast , a promising host for lignocellulosic bioethanol production, is unable to metabolize xylose. In attempts to confer xylose utilization ability in , a number of xylose isomerase (XI) genes have been expressed heterologously in this yeast. Although several of these XI encoding genes were functionally expressed in , the need still exists for a strain with improved xylose utilization ability for use in the commercial production of bioethanol. Although currently much effort has been devoted to achieve the objective, one of the solutions is to search for a new XI gene that would confer superior xylose utilization in . Here, we searched for novel XI genes from the protists residing in the hindgut of the termite . Eight novel XI genes were obtained from a cDNA library, prepared from the protists of the hindgut, by PCR amplification using degenerated primers based on highly conserved regions of amino acid sequences of different XIs. Phylogenetic analysis classified these cloned XIs into two groups, one showed relatively high similarities to and the other was comparatively similar to . The growth rate and the xylose consumption rate of the strain expressing the novel XI, which exhibited highest XI activity among the eight XIs, were superior to those exhibited by the strain expressing the XI gene from sp. E2. Substitution of the asparagine residue at position 337 of the novel XI with a cysteine further improved the xylose utilization ability of the yeast strain. Interestingly, introducing point mutations in the corresponding asparagine residues in XIs originated from other organisms, such as sp. E2 or , similarly improved xylose utilization in . A novel XI gene conferring superior xylose utilization in was successfully isolated from the protists in the termite hindgut. Isolation of this XI gene and identification of the point mutation described in this study might contribute to improving the productivity of industrial bioethanol.