Structure-function relationships of LamB protein, an outer membrane protein of Escherichia coli, will be investigated to elucidate its role in membrane transport. Mutant LamB proteins will be purified from strains of E. coli carrying missense mutations in the lamB gene. Transport activity of the mutant LamB proteins will be examined in reconstituted liposomes. Determination of secondary structure of mutant and parent proteins will be based on circular dichroism spectra, and the presence and role of disulfhydryls will be investigated. The interaction of LamB protein with periplasmic maltose binding protein (MBP) will be probed by characterization of mutant proteins with decreased affinity for MBP and by effects of chemical modification of LamB protein on the interaction. Crystallization of LamB protein will be attempted using equilibrium dialysis techniqes, for knowledge of its detailed three dimensional structure will come ultimately from Xray analysis. The LamB protein is part of the maltose transport system, and facilitates the transport of maltose and maltodextrins across the outer membrane. An early model of its mode of action suggested it forms a nonspecific pore across the outer membrane which gains specificity only when """"""""gated"""""""" by MBP. However, reconstitution studies of the purified LamB protein (in absence of MBP) revealed marked specificity of the transport process, in contrast to the nonspecific channel formed by porins. The goals of this work are to delineate the nature of transport through the Lamb protein channel and to understand the role of MBP in this transport. The results may be applicable to a number of other bacterial transport systems which contain periplasmic binding proteins, since the precise role of the binding proteins is still unknown. In addition, the work will contribute to a better description of the role of proteins in facilitated diffusion, a fundamental role of many membrane proteins that contributes to the selective permeability of membranes. Knowledge of the basic structure/fucntion relationships of membrane proteins can be expected to contribute to our understanding of many disease processes. In addition, characterization of the outer membrane of Gram negative bacteria may further our understanding of bacterial infections and antibiotic resistance among bacteria.
|Luckey, M; Ling, R; Dose, A et al. (1991) Role of a disulfide bond in the thermal stability of the LamB protein trimer in Escherichia coli outer membrane. J Biol Chem 266:1866-71|