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J. Bacteriol. doi:10.1128/JB.01620-06
Copyright (c) 2007, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Properties of R-citramalyl-CoA Lyase and Its Role in the Autotrophic 3-Hydroxypropionate Cycle of Chloroflexus aurantiacus

Mikrobiologie, Institut Biologie II, Universität Freiburg, Freiburg, Germany

^* To whom correspondence should be addressed. Email: georg.fuchs{at}biologie.uni-freiburg.de.

	Abstract

The autotrophic CO₂ fixation pathway in Chloroflexus aurantiacus (3-hydroxypropionate cycle) results in the fixation of two molecules of bicarbonate into one molecule of glyoxylate. Glyoxylate conversion to the CO₂ acceptor molecule acetyl-CoA requires condensation with propionyl-CoA (derived from one molecule of acetyl-CoA and one molecule of CO₂) to -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 cell carbon biosynthesis. Both S- and R-citramalyl-CoA lyases were up-regulated several-fold during autotrophic growth. S-Citramalyl-CoA lyase activity was found to be due to L-malyl-CoA lyase/-methylmalyl-CoA lyase. This promiscuous enzyme is involved in the CO₂ fixation pathway, forms acetyl-CoA and glyoxylate from L-malyl-CoA, and condenses glyoxylate with propionyl-CoA to -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 Mg² or Mn²⁺ 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.

Similar genes involved in CO₂ fixation of C. aurantiacus, including a R-citramalyl-CoA lyase gene, were found in Roseiflexus sp. suggesting the operation of the 3-hydroxypropionate cycle in this bacterium. An incomplete set of genes was found in aerobic phototrophic bacteria and in the -proteobacterium Congregibacter litoralis. This may indicate that part of the reactions may be involved in a different metabolic process.

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