Molecular cloning of two different cDNAs for maize acetyl CoA carboxylase

Anthony R. Ashton, Colin L.D. Jenkins, Paul R. Whitfeld

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

Acetyl CoA carboxylase (EC 6.4.1.2) in plants is a chloroplast-localized, biotin-containing enzyme that catalyses the carboxylation of acetyl CoA to malonyl CoA, the first committed step of the fatty acid biosynthesis pathway. Acetyl CoA carboxylase is the target site for the monocotyledon-specific aryloxy-phenoxypropionate and cyclohexanedione groups of herbicides. We have purifed a herbicide-sensitive acetyl CoA carboxylase from maize leaves to homogeneity (specific activity 7 μmol min-1 mg-1), separating it during the purification from a minor herbicide-resistant acetyl CoA carboxylase. The purified enzyme is a dimer of 230 kDa subunits. Antibodies raised to the purified acetyl CoA carboxylase detected three cross-reacting clones in a maize leaf cDNA expression library, each having an insert of 4-4.5 kb. Restriction analysis and sequencing showed that the cDNAs were derived from two different transcripts. Comparison of the deduced amino acid sequences with those of chicken and yeast acetyl CoA carboxylases confirmed that both types encoded acetyl CoA carboxylase, corresponding to the C-terminal half of the enzyme. The overall identity of the maize and chicken sequences was 37% (58% similarity) but for some shorter regions was much higher. Analysis of six other acetyl CoA carboxylase clones recovered from the maize cDNA library showed four belonged to one type and two to the other. The nucleotide sequence similarity between the two types of cDNA was approximately 95% in the coding region but considerably less in the 3′-untranslated region. Northern blot analysis of maize RNA showed a single band of 8.2-8.5 kb for acetyl CoA carboxylase mRNA. Southern blot hybridisations indicated that there are probably no more than two genes in maize for acetyl CoA carboxylase. The possible significance of two different cDNAs for acetyl CoA carboxylase is discussed.

Original languageEnglish
Pages (from-to)35-49
Number of pages15
JournalPlant Molecular Biology
Volume24
Issue number1
DOIs
Publication statusPublished - Jan 1994
Externally publishedYes

Keywords

  • acetyl CoA carboxylase
  • cDNA Cloning
  • herbicide
  • nucleotide sequence
  • purification
  • Zea mays

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