Properties of R-citramalyl-coenzyme A lyase and its role in the autotrophic 3-hydroxypropionate cycle of Chloroflexus aurantiacus.
The autotrophic CO(2) fixation pathway (3-hydroxypropionate cycle) in Chloroflexus aurantiacus results in the fixation of two molecules of bicarbonate into one molecule of glyoxylate. Glyoxylate conversion to the CO(2) acceptor molecule acetyl-coenzyme A (CoA) requires condensation with propionyl-CoA (derived from one molecule of acetyl-CoA and one molecule of CO(2)) to beta-methylmalyl-CoA, which is converted to citramalyl-CoA. Extracts of autotrophically grown cells contained both S- and R-citramalyl-CoA lyase activities, which formed acetyl-CoA and pyruvate. Pyruvate is taken out of the cycle and used for cellular carbon biosynthesis. Both the S- and R-citramalyl-CoA lyases were up-regulated severalfold during autotrophic growth. S-Citramalyl-CoA lyase activity was found to be due to l-malyl-CoA lyase/beta-methylmalyl-CoA lyase. This promiscuous enzyme is involved in the CO(2) fixation pathway, forms acetyl-CoA and glyoxylate from l-malyl-CoA, and condenses glyoxylate with propionyl-CoA to beta-methylmalyl-CoA. R-Citramalyl-CoA lyase was further studied. Its putative gene was expressed and the recombinant protein was purified. This new enzyme belongs to the 3-hydroxy-3-methylglutaryl-CoA lyase family and is a homodimer with 34-kDa subunits that was 10-fold stimulated by adding Mg(2) or Mn(2+) ions and dithioerythritol. The up-regulation under autotrophic conditions suggests that the enzyme functions in the ultimate step of the acetyl-CoA regeneration route in C. aurantiacus. Genes similar to those involved in CO(2) fixation in C. aurantiacus, including an R-citramalyl-CoA lyase gene, were found in Roseiflexus sp., suggesting the operation of the 3-hydroxypropionate cycle in this bacterium. Incomplete sets of genes were found in aerobic phototrophic bacteria and in the gamma-proteobacterium Congregibacter litoralis. This may indicate that part of the reactions may be involved in a different metabolic process.
Friedmann
Silke
S
Mikrobiologie, Institut Biologie II, Schänzlestr. 1, D-79104 Freiburg, Germany.
Alber
Birgit E
BE
Fuchs
Georg
G
D000595:Amino Acid Sequence; D002245:Carbon Dioxide / Q000378:metabolism; D019755:Carbon-Carbon Lyases / Q000235:genetics / Q000378:metabolism*; D041861:Chloroflexus / Q000201:enzymology*; D020563:Gammaproteobacteria / Q000201:enzymology; D006038:Glyoxylates / Q000378:metabolism; D007700:Kinetics; D019344:Lactic Acid / Q000031:analogs & derivatives* / Q000378:metabolism; D008969:Molecular Sequence Data; D010802:Phylogeny; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D006038:Glyoxylates; D011994:Recombinant Proteins; D002245:Carbon Dioxide; D019344:Lactic Acid; C031601:hydracrylic acid; D019755:Carbon-Carbon Lyases; C519717:R-citramalyl-coenzyme A lyase, Chloroflexus aurantiacus; C031150:glyoxylic acid