title: Glycerol dehydrogenase plays a dual role in glycerol metabolism and 2,3-butanediol formation in Klebsiella pneumoniae.
abstract: Glycerol dehydrogenase (GDH) is an important polyol dehydrogenase for glycerol metabolism in diverse microorganisms and for value-added utilization of glycerol in the industry. Two GDHs from Klebsiella pneumoniae, DhaD and GldA, were expressed in Escherichia coli, purified and characterized for substrate specificity and kinetic parameters. Both DhaD and GldA could catalyze the interconversion of (3R)-acetoin/(2R,3R)-2,3-butanediol or (3S)-acetoin/meso-2,3-butanediol, in addition to glycerol oxidation. Although purified GldA appeared more active than DhaD, in vivo inactivation and quantitation of their respective mRNAs indicate that dhaD is highly induced by glycerol and plays a dual role in glycerol metabolism and 2,3-butanediol formation. Complementation in K. pneumoniae further confirmed the dual role of DhaD. Promiscuity of DhaD may have vital physiological consequences for K. pneumoniae growing on glycerol, which include balancing the intracellular NADH/NAD(+) ratio, preventing acidification, and storing carbon and energy. According to the kinetic response of DhaD to modified NADH concentrations, DhaD appears to show positive homotropic interaction with NADH, suggesting that the physiological role could be regulated by intracellular NADH levels. The co-existence of two functional GDH enzymes might be due to a gene duplication event. We propose that whereas DhaD is specialized for glycerol utilization, GldA plays a role in backup compensation and can turn into a more proficient catalyst to promote a survival advantage to the organism. Revelation of the dual role of DhaD could further the understanding of mechanisms responsible for enzyme evolution through promiscuity, and guide metabolic engineering methods of glycerol metabolism.
authors.affiliation: From the State Key Laboratory of Microbial Metabolism, and School of Life Sciences & Biotechnology, Shanghai Jiao Tong University, Shanghai 200240, People's Republic of China.
authors.forename: Yu
authors.initials: Y
authors.lastname: Wang
authors.forename: Fei
authors.initials: F
authors.lastname: Tao
authors.forename: Ping
authors.initials: P
authors.lastname: Xu
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D005990:Glycerol / Q000737:chemistry* / Q000378:metabolism; D007711:Klebsiella pneumoniae / Q000201:enzymology* / Q000235:genetics; D050296:Microbial Viability; D009243:NAD / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D010084:Oxidation-Reduction; D013401:Sugar Alcohol Dehydrogenases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; C109038:GldA protein, bacteria; D009243:NAD; D013401:Sugar Alcohol Dehydrogenases; C020672:glycerol dehydrogenase; D005990:Glycerol
keywords: 2,3-Butanediol; Bacterial Metabolism; Dehydrogenase; Enzyme Kinetics; Enzyme Promiscuity; Glycerol; Glycerol Dehydrogenase; Klebsiella pneumonia; Protein Evolution