Mutations that affect ligand binding to the Escherichia coli aspartate receptor - Implications for transmembrane signaling

Abstract

Three arginine residues of the binding site of the Escherchia coli aspartate receptor contribute to its high affinity for aspartate (K-d similar to3 mum). Site-directed mutations at residue 64 had the greatest effect on aspartate binding. No residue could substitute for the native arginine; all changes resulted in an apparent K-d of similar to 35 mM. These mutations had little impact on maltose responses. At residue Arg-69, a lysine substitution was least disruptive, conferring an apparent K-d of 0.3 mM for aspartate, Results obtained for an alanine mutant were similar to those with cysteine and histidine mutants (K-d similar to5 mM) indicating that side chain size was not an important factor here. Proline and aspartate caused more severe defects, presumably for reasons related to conformation and charge. The impact of residue 69 mutations on the maltose response was small. Mutations at Arg-73 had similar effects on aspartate binding (K-d 0.3-7 mm) but more severe consequences for maltose responses. Larger side chains resulted in the best aspartate binding, implying steric considerations are important here. Signaling in the mutant proteins was surprisingly robust. Given aspartate binding, signaling occurred with essentially wild-type efficiency. These results were evaluated in the context of available structural data.

Published in

Journal of Biological Chemistry
2001, volume: 276, number: 4, pages: 2808-2815
Publisher: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC

SLU Authors

• Björkman, Malin

  • Department of Molecular Biology, Swedish University of Agricultural Sciences
    

• Sandgren, Mats

  • Uppsala University

• Mowbray, Sherry

  • Department of Molecular Biology, Swedish University of Agricultural Sciences
    

UKÄ Subject classification

Microbiology
Biochemistry and Molecular Biology

Publication identifier

• DOI: https://doi.org/10.1074/jbc.M009593200

Permanent link to this page (URI)

https://res.slu.se/id/publ/43055