Amino acid substitution in the lactose carrier protein with the use of amber suppressors

A. M. Huang, J. I. Lee, S. C. King, T. H. Wilson

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Abstract

Five lacY mutants with amber stop codons at known positions were each placed into 12 different suppressor strains. The 60 amino acid substitutions obtained in this manner were tested for growth on lactose-minimal medium plates and for transport of lactose, melibiose, and thiomethylgalactoside. Most of the amino acid substitutions in the regions of the putative loops (between transmembrane α helices) resulted in a reasonable growth rate on lactose with moderate-to-good transport activity. In one strain (glycine substituted for Trp-10), abnormal sugar recognition was found. The substitution of proline for Trp-33 (in the region of the first α helix) showed no activity, while four additional substitutions (lysine, leucine, cysteine, and glutamic acid) showed low activity. Altered sugar specificity was observed when Trp-33 was replaced by serine, glutamine, tyrosine, alanine, histidine, or phenylalanine. It is concluded that Trp-33 may be involved directly or indirectly in sugar recognition.

Original languageEnglish (US)
Pages (from-to)5436-5441
Number of pages6
JournalJournal of bacteriology
Volume174
Issue number16
DOIs
StatePublished - Jan 1 1992

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ASJC Scopus subject areas

  • Microbiology
  • Molecular Biology

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