Protein Science, Vol 6, Issue 3 628-636, Copyright © 1997 by Cold Spring Harbor Laboratory Press

ARTICLE

Residues in the {alpha} helix 7 of the bacterial maltose binding protein which are important in interactions with the Mal FGK(2) complex

S. SZMELCMAN, N. SASSOON and M. HOFNUNG
Unite de Programmation Moleculaire & Toxicologie Genetique--CNRS Ura 1444 Institut Pasteur, 25 rue du Dr Roux, 75015 Paris, France

The periplasmic maltose binding protein, MalE, is a major element in maltose transport and in chemotaxis towards this sugar. Previous genetic analysis of the MalE protein revealed functional domains involved in transport and chemotactic functions. Among them the surface located {alpha} helix 7, which is part of the C-lobe, one of the two lobes forming the three dimensional structure of MalE. Small deletions in this region abolished maltose transport, although maintaining wild-type affinity and specificity as well as a normal chemoreceptor function. It was suggested that {alpha} helix 7 may be implicated in interactions between the maltose binding protein and the membrane-bound protein complex (Duplay P, Szmelcman S. 1987. Silent and functional changes in the periplasmic maltose binding protein of Escherichia coli K12. II. Chemotaxis towards maltose. J Mol Biol 194:675-678; Duplay P, Szmelcman S, Bedouelle H, Hofnung M. 1987. Silent and functional changes in the periplasmic maltose binding protein of Escherichia coli K12. I: Transport of maltose. J Mol Biol 194:663-673). In this study, we submitted a region of 14 residues--Asp 207 to Gly 220--encompassing {alpha} helix 7, to genetic analysis by oligonucleotide mediated random mutagenesis. Out of 127 identified mutations, twelve single and five double mutants with normal affinities towards maltose were selected for further investigation. Two types of mutations were characterized, silent mutations that did not affect maltose transport and mutations that heavily impaired transport kinetics, even though the maltose binding capacity of the mutant proteins remained normal. Three substitutions at Tyr 210 (Y210S, Y210L, Y210N) drastically reduced maltose transport. One substitution at Ala 213 (A213I) and one substitution at Glu 214 (E214K) also impaired transport. These three identified residues, Tyr 210, Ala 213, and Glu 214, which are constituents of {alpha} helix 7, therefore seem to play some important role in maltose transport, most probably in a productive interaction between the MalE protein and the membrane bound MalFGK(2) complex.
CiteULike    Connotea    Del.icio.us    Digg    Reddit    Technorati    What's this?

This article has been cited by other articles:

				A. L. Davidson, E. Dassa, C. Orelle, and J. Chen Structure, Function, and Evolution of Bacterial ATP-Binding Cassette Systems Microbiol. Mol. Biol. Rev., June 1, 2008; 72(2): 317 - 364. [Abstract] [Full Text] [PDF]

				E. L. Borths, K. P. Locher, A. T. Lee, and D. C. Rees The structure of Escherichia coli BtuF and binding to its cognate ATP binding cassette transporter PNAS, December 24, 2002; 99(26): 16642 - 16647. [Abstract] [Full Text] [PDF]

				J. D. Fox, R. B. Kapust, and D. S. Waugh Single amino acid substitutions on the surface of Escherichia coli maltose-binding protein can have a profound impact on the solubility of fusion proteins Protein Sci., March 1, 2001; 10(3): 622 - 630. [Abstract] [Full Text]

				W. Boos and H. Shuman Maltose/Maltodextrin System of Escherichia coli: Transport, Metabolism, and Regulation Microbiol. Mol. Biol. Rev., March 1, 1998; 62(1): 204 - 229. [Abstract] [Full Text] [PDF]