Complex dynamics and spatial distributions of membrane proteins in the bacterial cell envelope

For many proteins and multiprotein structures, proper biological function requires a dynamic, non-uniform spatial distribution. Advanced techniques for observing protein localization in live bacteria show that the distributions are dynamic. For technical reasons, most such techniques have not been applied to outer membrane proteins in Gram-negative bacteria. Here, we have determined the distributions and dynamics of two Escherichia coli integral membrane proteins---LamB and MdfA. We have developed two novel live-cell imaging techniques to observe the surface distribution of LamB, an abundant integral outer membrane protein in E. coli responsible for maltose uptake and for attachment of bacteriophage lambda. Using fluorescently labeled bacteriophage lambda tails, we quantitatively described the spatial distribution and dynamic movement of LamB in the outer membrane. LamB accumulated in spiral patterns. The distribution depended on cell length and changed rapidly. The majority of the protein diffused along spirals extending across the cell body. Tracking single particles, we found that there are two populations of LamB---one shows very restricted diffusion and the other shows greater mobility. The presence of two populations recalls the partitioning of eukaryotic membrane proteins between "mobile" and "immobile" populations.; Preliminary experiments have shown that the multidrug-resistant efflux pump, MdfA, also has a dynamic, non-uniform distribution in the membrane. MdfA, an integral inner membrane protein, was observed dispersed along the membrane, and some protein accumulations appeared to migrate across the surface. The protein distributions of MdfA changed moderately in the presence of actively exported antibiotics, and strikingly, our fusion protein, MdfA-GFP, was expressed in the presence of antibiotics, even in the absence of induction. This suggested that MdfA expression might be regulated by mRNA or protein stability mechanisms that are sensitive to the presence of its ligands. Through these studies, we have begun to probe the distributions and dynamics of integral membrane proteins in bacteria and have launched new directions for future inquiries in bacterial cell biology.