Pyruvate formate-lyase and a novel route of eukaryotic ATP synthesis in Chlamydomonas mitochondria.
Pyruvate formate-lyase (PFL) catalyzes the non-oxidative conversion of pyruvate to formate and acetyl-CoA. PFL and its activating enzyme (PFL-AE) are common among strict anaerobic and microaerophilic prokaryotes but are very rare among eukaryotes. In a proteome survey of isolated Chlamydomonas reinhardtii mitochondria, we found several PFL-specific peptides leading to the identification of cDNAs for PFL and PFL-AE, establishing the existence of a PFL system in this photosynthetic algae. Anaerobiosis and darkness led to increased PFL transcripts but had little effect on protein levels, as determined with antiserum raised against C. reinhardtii PFL. Protein blots revealed the occurrence of PFL in both chloroplast and mitochondria purified from aerobically grown cells. Mass spectrometry sequencing of C. reinhardtii mitochondrial proteins, furthermore, identified peptides for phosphotransacetylase and acetate kinase. The phosphotransacetylase-acetate kinase pathway is a common route of ATP synthesis or acetate assimilation among prokaryotes but is novel among eukaryotes. In addition to PFL and pyruvate dehydrogenase, the algae also expresses pyruvate:ferredoxin oxidoreductase and bifunctional aldehyde/alcohol dehydrogenase. Among eukaryotes, the oxygen producer C. reinhardtii has the broadest repertoire of pyruvate-, ethanol-, and acetate-metabolizing enzymes described to date, many of which were previously viewed as specific to anaerobic eukaryotic lineages.
Atteia
Ariane
A
Institute of Botany, University of Düsseldorf, 40225 Düsseldorf, Germany. aatteia@cea.fr
van Lis
Robert
R
Gelius-Dietrich
Gabriel
G
Adrait
Annie
A
Garin
Jérôme
J
Joyard
Jacques
J
Rolland
Norbert
N
Martin
William
W
D000115:Acetylesterase / Q000378:metabolism; D000123:Acetyltransferases / Q000502:physiology*; D000255:Adenosine Triphosphate / Q000737:chemistry*; D000595:Amino Acid Sequence; D000818:Animals; D016825:Chlamydomonas reinhardtii / Q000378:metabolism*; D002736:Chloroplasts / Q000378:metabolism; D018076:DNA, Complementary / Q000378:metabolism; D020224:Expressed Sequence Tags; D015723:Gene Library; D015151:Immunoblotting; D013058:Mass Spectrometry; D008928:Mitochondria / Q000378:metabolism*; D008954:Models, Biological; D008956:Models, Chemical; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry; D010802:Phylogeny; D040901:Proteomics / Q000379:methods; D011768:Pyruvate Dehydrogenase Complex / Q000737:chemistry; D050743:Pyruvate Synthase / Q000378:metabolism; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D018076:DNA, Complementary; D010455:Peptides; D011768:Pyruvate Dehydrogenase Complex; D012333:RNA, Messenger; D012313:RNA; D000255:Adenosine Triphosphate; D050743:Pyruvate Synthase; D000123:Acetyltransferases; C022560:formate C-acetyltransferase; D000115:Acetylesterase