title: Malonyl-coenzyme A reductase in the modified 3-hydroxypropionate cycle for autotrophic carbon fixation in archaeal Metallosphaera and Sulfolobus spp.
abstract: Autotrophic members of the Sulfolobales (Crenarchaeota) contain acetyl-coenzyme A (CoA)/propionyl-CoA carboxylase as the CO2 fixation enzyme and use a modified 3-hydroxypropionate cycle to assimilate CO2 into cell material. In this central metabolic pathway malonyl-CoA, the product of acetyl-CoA carboxylation, is further reduced to 3-hydroxypropionate. Extracts of Metallosphaera sedula contained NADPH-specific malonyl-CoA reductase activity that was 10-fold up-regulated under autotrophic growth conditions. Malonyl-CoA reductase was partially purified and studied. Based on N-terminal amino acid sequencing the corresponding gene was identified in the genome of the closely related crenarchaeum Sulfolobus tokodaii. The Sulfolobus gene was cloned and heterologously expressed in Escherichia coli, and the recombinant protein was purified and studied. The enzyme catalyzes the following reaction: malonyl-CoA + NADPH + H+ --> malonate-semialdehyde + CoA + NADP+. In its native state it is associated with small RNA. Its activity was stimulated by Mg2+ and thiols and inactivated by thiol-blocking agents, suggesting the existence of a cysteine adduct in the course of the catalytic cycle. The enzyme was specific for NADPH (Km = 25 microM) and malonyl-CoA (Km = 40 microM). Malonyl-CoA reductase has 38% amino acid sequence identity to aspartate-semialdehyde dehydrogenase, suggesting a common ancestor for both proteins. It does not exhibit any significant similarity with malonyl-CoA reductase from Chloroflexus aurantiacus. This shows that the autotrophic pathway in Chloroflexus and Sulfolobaceae has evolved convergently and that these taxonomic groups have recruited different genes to bring about similar metabolic processes.
authors.affiliation: Mikrobiologie, Institut Biologie II, Schänzlestr. 1, D-79104 Freiburg, Germany.
authors.forename: Birgit
authors.initials: B
authors.lastname: Alber
authors.forename: Marc
authors.initials: M
authors.lastname: Olinger
authors.forename: Annika
authors.initials: A
authors.lastname: Rieder
authors.forename: Daniel
authors.initials: D
authors.lastname: Kockelkorn
authors.forename: Björn
authors.initials: B
authors.lastname: Jobst
authors.forename: Michael
authors.initials: M
authors.lastname: Hügler
authors.forename: Georg
authors.initials: G
authors.lastname: Fuchs
mesh_terms: D000595:Amino Acid Sequence; D002245:Carbon Dioxide / Q000378:metabolism; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D019344:Lactic Acid / Q000031:analogs & derivatives* / Q000378:metabolism; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D016943:Sulfolobaceae / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D016944:Sulfolobus / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development
chemical_list: D011994:Recombinant Proteins; D002245:Carbon Dioxide; D019344:Lactic Acid; D009249:NADP; C031601:hydracrylic acid; D010088:Oxidoreductases; C454539:malonyl-Coa reductase