Identification of an l-alanine export system in Escherichia coli and isolation and characterization of export-deficient mutants

Hatsuhiro Hori, Tasuke Ando, Emiko Isogai, Hiroshi Yoneyama, Ryoichi Katsumata

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10 Citations (Scopus)

Abstract

An Escherichia coli strain that exhibits a double auxotrophy for l-alanine and d-alanine was constructed. During growth in the presence of the dipeptide l-alanyl-l-alanine (Ala-Ala), this was fully consumed with concomitant extracellular accumulation of l-alanine in a twofold molar concentration compared with the dipeptide. This finding indicates that the strain not only can hardly degrade l-alanine but has an export system(s) for l-alanine. To obtain access to the system, we chemically mutagenized the l-alanine-nonmetabolizing strain and isolated mutants with increased Ala-Ala sensitivity. Two such mutants accumulated l-alanine up to 150-190mM in the cytoplasm with a reduced rate of l-alanine export relative to the parent strain in the presence of Ala-Ala. Furthermore, when chloramphenicol was added together with Ala-Ala, the parent strain accumulated l-alanine in the cytoplasm to a level similar to that observed in the mutants in the absence of chloramphenicol. In contrast, the intracellular l-alanine level in the mutants did not change irrespective of chloramphenicol treatment. From these results, we conclude that E. coli has an inducible l-alanine export carrier, together with a second, as yet unidentified, mechanism of alanine export.

Original languageEnglish
Pages (from-to)83-89
Number of pages7
JournalFEMS Microbiology Letters
Volume316
Issue number2
DOIs
Publication statusPublished - 2011 Mar

Keywords

  • Alanine auxotroph
  • Escherichia coli
  • L-alanine export
  • Peptide sensitivity

ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology
  • Genetics

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