Isolation and characterization of ECT1 gene encoding CTP: phosphoethanolamine cytidylyltransferase of Saccharomyces cerevisiae

J Biochem. 1996 Nov;120(5):1040-7. doi: 10.1093/oxfordjournals.jbchem.a021497.

Abstract

Saccharomyces cerevisiae mutants that were unable to utilize extracellular ethanolamine for phosphatidylethanolamine synthesis were isolated. Two of them carried recessive chromosomal mutations in a same gene and were defective in CTP:phosphoethanolamine cytidylyltransferase (ECT) activity in vitro (Ect-). In an Ect- mutant that also carried the cho1 mutation, phosphatidylethanolamine accounted for less than 2% of total phospholipids, suggesting the importance of ECT in phosphatidylethanolamine synthesis. By screening a genomic library on a low copy number vector, three complementary clones of different size were isolated. A 2.8-kb common DNA region carried an open reading frame (ORF) of 969 bp in length, of which a truncated from failed to complement the Ect- mutation. This ORF was identical to the previously isolated MUQ1 gene of unknown function. Its deduced amino acid sequence had significant similarity to CTP: phosphocholine cytidylyl-transferases of yeast and rat. The entire ORF, when combined with the glutathione S-transferase gene and expressed in Escherichia coli, exhibited ECT activity. These results indicate that the cloned gene encodes a catalytic subunit of ECT of S. cerevisiae.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Base Sequence
  • CHO Cells
  • Cricetinae
  • Molecular Sequence Data
  • Mutagenesis, Site-Directed
  • Nucleotidyltransferases / chemistry
  • Nucleotidyltransferases / genetics*
  • RNA Nucleotidyltransferases
  • Rats
  • Saccharomyces cerevisiae / enzymology*
  • Sequence Alignment

Substances

  • Nucleotidyltransferases
  • RNA Nucleotidyltransferases
  • ECT1 protein, S cerevisiae
  • Ethanolamine-phosphate cytidylyltransferase

Associated data

  • GENBANK/D50644