Isolation of lactose permease mutants which recognize arabinose

Visala Chepuri Goswitz, Robert J Brooker

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

In the present study lactose permease mutants were isolated which recognize the monosaccharide, L-arabinose. Although the wild-type permease exhibits a poor recognition for L-arabinose, seven independent mutants were identified by their ability to grow on L-arabinose minimal plates. When subjected to DNA sequencing, it was found that all seven of these mutants were single-site mutations in which alanine 177 was changed to valine. The wild type and valine 177 mutant were then analyzed with regard to their abilities to recognize and transport monosaccharides and disaccharides. Free L-arabinose was shown to competitively inhibit [14C]-lactose transport yielding aKi value of 121 mM for the Val177 mutant and a much higher value of 320 mM for the wild-type. Among several monosaccharides, D-glucose as well as L-arabinose inhibited lactose transport in the Val177 mutant to a significantly greater extent, while D-arabinose and D-xylose only caused a slight inhibition. On the other hand, kinetic studies with sugars which are normally recognized by the wild-type permease such as [14C]-galactose and [14C]-lactose revealed that the Val177 mutant and wild-type strains had similar transport characteristics for these two sugars. Overall, these results are consistent with the notion that the Val177 substitution causes an enhanced recognition for particular sugars (i.e. L-arabinose) but does not universally affect the recognition and unidirectional transport for all sugars. This idea is further supported by the observation that site-directed mutants containing isoleucine, leucine, phenylalanine, or proline at position 177 also were found to possess an enhanced recognition for L-arabinose.

Original languageEnglish (US)
Pages (from-to)61-70
Number of pages10
JournalMolecular Membrane Biology
Volume10
Issue number1
DOIs
StatePublished - Jan 1 1993

Fingerprint

Arabinose
Sugars
Monosaccharides
Lactose
Membrane Transport Proteins
Valine
lactose permease
Isoleucine
Disaccharides
Xylose
Phenylalanine
Galactose
DNA Sequence Analysis
Proline
Leucine
Alanine
Substitution reactions
Glucose
Mutation
Kinetics

Keywords

  • Arabinose
  • Lactose permease
  • Membrane protein
  • Substrate specificity

Cite this

Isolation of lactose permease mutants which recognize arabinose. / Goswitz, Visala Chepuri; Brooker, Robert J.

In: Molecular Membrane Biology, Vol. 10, No. 1, 01.01.1993, p. 61-70.

Research output: Contribution to journalArticle

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