title: Characterization of 5-chloro-5-deoxy-D-ribose 1-dehydrogenase in chloroethylmalonyl coenzyme A biosynthesis: substrate and reaction profiling.
abstract: SalM is a short-chain dehydrogenase/reductase enzyme from the marine actinomycete Salinispora tropica that is involved in the biosynthesis of chloroethylmalonyl-CoA, a novel halogenated polyketide synthase extender unit of the proteasome inhibitor salinosporamide A. SalM was heterologously overexpressed in Escherichia coli and characterized in vitro for its substrate specificity, kinetics, and reaction profile. A sensitive real-time (13)C NMR assay was developed to visualize the oxidation of 5-chloro-5-deoxy-D-ribose to 5-chloro-5-deoxy-D-ribono-γ-lactone in an NAD(+)-dependent reaction, followed by spontaneous lactone hydrolysis to 5-chloro-5-deoxy-D-ribonate. Although short-chain dehydrogenase/reductase enzymes are widely regarded as metal-independent, a strong divalent metal cation dependence for Mg(2+), Ca(2+), or Mn(2+) was observed with SalM. Oxidative activity was also measured with the alternative substrates D-erythrose and D-ribose, making SalM the first reported stereospecific non-phosphorylative ribose 1-dehydrogenase.
authors.affiliation: Center of Marine Biotechnology and Biomedicine, Scripps Institution of Oceanography, University of California at San Diego, La Jolla, California 92093, USA.
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Kale
authors.forename: Ryan P
authors.initials: RP
authors.lastname: McGlinchey
authors.forename: Bradley S
authors.initials: BS
authors.lastname: Moore
mesh_terms: D000429:Alcohol Oxidoreductases / Q000737:chemistry*; D001426:Bacterial Proteins / Q000737:chemistry*; D001483:Base Sequence; D002241:Carbohydrates / Q000737:chemistry; D002384:Catalysis; D002412:Cations; D003065:Coenzyme A / Q000737:chemistry*; D004798:Enzymes / Q000737:chemistry; D004926:Escherichia coli / Q000378:metabolism; D006094:Gram-Positive Bacteria / Q000378:metabolism*; D007700:Kinetics; D009682:Magnetic Resonance Spectroscopy / Q000379:methods; D008670:Metals / Q000737:chemistry; D008969:Molecular Sequence Data; D009154:Mutation; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D002241:Carbohydrates; D002412:Cations; D004798:Enzymes; D008670:Metals; C553401:5-chloro-5-deoxy-D-ribose 1-dehydrogenase, Salinispora tropica; D000429:Alcohol Oxidoreductases; D003065:Coenzyme A