Carnitine-dependent metabolic activities in Saccharomyces cerevisiae: three carnitine acetyltransferases are essential in a carnitine-dependent strain.
L-carnitine is required for the transfer of activated acyl-groups across intracellular membranes in eukaryotic organisms. In Saccharomyces cerevisiae, peroxisomal membranes are impermeable to acetyl-CoA, which is produced in the peroxisome when cells are grown on fatty acids as carbon source. In a reversible reaction catalysed by carnitine acetyltransferases (CATs), activated acetyl groups are transferred to carnitine to form acetylcarnitine which can be shuttled across membranes. Here we describe a mutant selection strategy that specifically selects for mutants affected in carnitine-dependent metabolic activities. Complementation of three of these mutants resulted in the cloning of three CAT encoding genes: CAT2, coding for the carnitine acetyltransferase associated with the peroxisomes and the mitochondria; YAT1, coding for the carnitine acetyltransferase, which is presumably associated with the outer mitochondrial membrane, and YER024w (YAT2), which encodes a third, previously unidentified carnitine acetyltransferase. The data also show that (a) L-carnitine and all three CATs are essential for growth on non-fermentable carbon sources in a strain with a disrupted CIT2 gene; (b) Yat2p contributes significantly to total CAT activity when cells are grown on ethanol; and that (c) the carnitine-dependent transfer of activated acetyl groups plays a more important role in cellular processes than previously realised.
Swiegers
J H
JH
Institute for Wine Biotechnology, University of Stellenbosch, Victoria Street, ZA-7600 Stellenbosch, South Africa.
Dippenaar
N
N
Pretorius
I S
IS
Bauer
F F
FF
D002331:Carnitine / Q000378:metabolism*; D002333:Carnitine Acyltransferases / Q000235:genetics / Q000378:metabolism*; D002332:Carnitine O-Acetyltransferase / Q000032:analysis; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D000431:Ethanol / Q000378:metabolism / Q000494:pharmacology; D005816:Genetic Complementation Test; D008565:Membrane Proteins / Q000032:analysis; D016296:Mutagenesis; D010957:Plasmids / Q000235:genetics; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D017422:Sequence Analysis, DNA
D004271:DNA, Fungal; D008565:Membrane Proteins; D029701:Saccharomyces cerevisiae Proteins; D000431:Ethanol; D002333:Carnitine Acyltransferases; D002332:Carnitine O-Acetyltransferase; C084974:YAT1 protein, S cerevisiae; D002331:Carnitine