Cloning and nucleotide sequence of the gene for protein X from Saccharomyces cerevisiae.
The gene encoding the protein X component of the pyruvate dehydrogenase complex from Saccharomyces cerevisiae has been cloned and sequenced. A 487-base fragment of yeast genomic DNA encoding the amino-terminal region of protein X was amplified by the polymerase chain reaction using synthetic oligonucleotide primers based on amino-terminal and internal amino acid sequences. This DNA fragment was used as a probe to select two genomic DNA restriction fragments, which were cloned and sequenced. A 2.1-kilobase insert contains the complete sequence of the protein X gene. This insert has an open reading frame of 1230 nucleotides encoding a presequence of 30 amino acid residues and a mature protein of 380 amino acid residues (Mr, 42,052). Hybridization analysis showed that there is a single copy of the protein X gene and that the size of the mRNA is approximately 1.5 kilobases. Comparison of the deduced amino acid sequences of yeast protein X and dihydrolipoamide acetyltransferase indicates that the two proteins evolved from a common ancestor. The amino-terminal part of protein X (residues 1-195) resembles the acetyltransferase, but the remainder is quite different. There is strong homology between protein X and the acetyltransferase in the amino-terminal region (residues 1-84) that corresponds to the putative lipoyl domain. Protein X lacks the highly conserved sequence His-Xaa-Xaa-Xaa-Asp-Gly near the carboxyl terminus, which is thought to be part of the active site of all dihydrolipoamide acyltransferases.
Behal
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RH
Clayton Foundation Biochemical Institute, University of Texas, Austin 78712.
Browning
K S
KS
Hall
T B
TB
Reed
L J
LJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics; D004259:DNA-Directed DNA Polymerase; D005656:Fungal Proteins / Q000235:genetics*; D005800:Genes, Fungal; D015698:Genomic Library; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010455:Peptides; D016133:Polymerase Chain Reaction; D011768:Pyruvate Dehydrogenase Complex / Q000235:genetics*; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D029701:Saccharomyces cerevisiae Proteins; D012689:Sequence Homology, Nucleic Acid
D004271:DNA, Fungal; D005656:Fungal Proteins; D015345:Oligonucleotide Probes; C489060:PDX1 protein, S cerevisiae; D010455:Peptides; D011768:Pyruvate Dehydrogenase Complex; D029701:Saccharomyces cerevisiae Proteins; D004259:DNA-Directed DNA Polymerase