title: Bacterial xylose isomerases from the mammal gut Bacteroidetes cluster function in Saccharomyces cerevisiae for effective xylose fermentation.
abstract: BACKGROUND
  Xylose isomerase (XI) catalyzes the conversion of xylose to xylulose, which is the key step for anaerobic ethanolic fermentation of xylose. Very few bacterial XIs can function actively in Saccharomyces cerevisiae. Here, we illustrate a group of XIs that would function for xylose fermentation in S. cerevisiae through phylogenetic analysis, recombinant yeast strain construction, and xylose fermentation.
  
  
  RESULTS
  Phylogenetic analysis of deposited XI sequences showed that XI evolutionary relationship was highly consistent with the bacterial taxonomic orders and quite a few functional XIs in S. cerevisiae were clustered with XIs from mammal gut Bacteroidetes group. An XI from Bacteroides valgutus in this cluster was actively expressed in S. cerevisiae with an activity comparable to the fungal XI from Piromyces sp. Two XI genes were isolated from the environmental metagenome and they were clustered with XIs from environmental Bacteroidetes group. These two XIs could not be expressed in yeast with activity. With the XI from B. valgutus expressed in S. cerevisiae, background yeast strains were optimized by pentose metabolizing pathway enhancement and adaptive evolution in xylose medium. Afterwards, more XIs from the mammal gut Bacteroidetes group, including those from B. vulgatus, Tannerella sp. 6_1_58FAA_CT1, Paraprevotella xylaniphila and Alistipes sp. HGB5, were individually transformed into S. cerevisiae. The known functional XI from Orpinomyces sp. ukk1, a mammal gut fungus, was used as the control. All the resulting recombinant yeast strains were able to ferment xylose. The respiration-deficient strains harboring B. vulgatus and Alistipes sp. HGB5 XI genes respectively obtained specific xylose consumption rate of 0.662 and 0.704 g xylose gcdw(-1) h(-1), and ethanol specific productivity of 0.277 and 0.283 g ethanol gcdw(-1) h(-1), much comparable to those obtained by the control strain carrying Orpinomyces sp. ukk1 XI gene.
  
  
  CONCLUSIONS
  This study demonstrated that XIs clustered in the mammal gut Bacteroidetes group were able to be expressed functionally in S. cerevisiae and background strain anaerobic adaptive evolution in xylose medium is essential for the screening of functional XIs. The methods outlined in this paper are instructive for the identification of novel XIs that are functional in S. cerevisiae.
authors.affiliation: School of Life Sciences and Chemical Technology, Ngee Ann Polytechnic, Singapore, Singapore. bingyin.peng@yahoo.com.
authors.forename: Bingyin
authors.initials: B
authors.lastname: Peng
authors.affiliation: School of Life Sciences and Chemical Technology, Ngee Ann Polytechnic, Singapore, Singapore. Higle945@gmail.com.|School of Chemical Engineering and Pharmacy, Key Laboratory for Green Chemical Process of Ministry of Education, Wuhan Institute of Technology, Wuhan, 430073, Peoples Republic of China. Higle945@gmail.com.
authors.forename: Shuangcheng
authors.initials: S
authors.lastname: Huang
authors.affiliation: School of Life Sciences and Chemical Technology, Ngee Ann Polytechnic, Singapore, Singapore. lti3@np.edu.sg.|School of Chemical Engineering and Pharmacy, Key Laboratory for Green Chemical Process of Ministry of Education, Wuhan Institute of Technology, Wuhan, 430073, Peoples Republic of China. lti3@np.edu.sg.
authors.forename: Tingting
authors.initials: T
authors.lastname: Liu
authors.affiliation: School of Life Sciences and Chemical Technology, Ngee Ann Polytechnic, Singapore, Singapore. gan2@np.edu.sg.
authors.forename: Anli
authors.initials: A
authors.lastname: Geng
mesh_terms: D019747:Aldose-Ketose Isomerases / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D041963:Bacteroidetes / Q000378:metabolism*; D005285:Fermentation; D005818:Genetic Engineering / Q000379:methods*; D008969:Molecular Sequence Data; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism
chemical_list: D019747:Aldose-Ketose Isomerases; C023298:xylose isomerase