The LamB protein, which is the receptor for bacteriophage lambda in the outer membrane of Escherichia coli, has been shown to form a transmembrane channel for uptake of maltose and maltodextrins. Purified LamB protein reconstituted in liposomes has been shown to discriminate strongly on the basis of saccharide configuration. The specificity appears to be conferred by one or more weak binding sites for the maltodextrin substrates. The proposed experiments will use a variety of biochemical and chemical techniques to investigate the nature of this transport channel. Chemical modifications and affinity labeling will be used to define portions of the protein which are involved in transport, pinpointing affected residues by HPLC mapping. The role or the periplasmic maltose binding protein (MP) will be studied by purification and characterization of mutant LamB proteins defective in binding MBP. Reconstitution studies and electron spin resonance experiment wil examine LamB protein's specific requirement for and interaction with phospholipid. Finally, new approaches to define the conformation of saccharides inside the channel will be developed using high resolution nuclear magnetic resonance. In carrying out passive transport across the outer membrane the LamB protein performs two of the steps necessary for more complex active transport processes: substrate recognition and solute translocation. Thus detailed molecular understanding of its transport mechanism should facilitate investigation of more complicated transport systems. In addition, the work will contribute to a better description of the role of proteins in facilitated diffusion, a fundamental role of other membrane proteins that contributes to the selective permeability of many membranes. Knowledge of the basic structure/function 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 infection and antibiotic resistance among bacteria.
|Ling, R; Luckey, M (1994) Use of single-cysteine mutants to probe the location of the disulfide bond in LamB protein from Escherichia coli. Biochem Biophys Res Commun 201:242-7|
|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|