Robert M. Weis, Department of Chemistry, University of Massachusetts, Amherst, MA 01003-9335
Physical & Biological Chemistry
Properties and processes of cell membrane and model membrane systems.
1. Mechanisms of Transmembrane Signaling through Cooperative Signaling Arrays. The chemosensory system in E. coli detects signals through a membrane array of transmembrane receptors and two associated signaling proteins: an adaptor protein (CheW) and the histidine kinase (CheA). The pathway creates a short term memory that allows a bacterium to compare the current concentrations of certain substances with those of the recent past (three to four seconds ago); this allows bacteria to bias swimming motion up gradients of favorable compounds (attractants), and down gradients of unfavorable compounds (repellents).
Chemotaxis, or rather chemotaxis-like pathways, are very widespread in prokaryotes. It now seems apparent many pathways don't regulate chemotaxis, yet the components common to all these pathways are the methyl-accepting chemotaxis proteins, CheW and CheA.
The components of the E. coli pathway and the pathway from Bacillus subtilis are depicted above for comparison.
Functional and structural studies both support the presence of a coopoerative receptor array in the membrane. Physically, these arrays look to be comprised of many hundreds to thousands of receptors, although the functional linkage -- accessed as cooperativity factors (Hill coefficients), indicate that the functional cluster is ~20 dimers; but this number is not known precisely. The cooperative inhibition of serine receptor/CheW/CheA signaling complexes (Li & Weis, 2000) are shown below. The cooperativity of inhibition increases with the level of covalent modification on the receptor (low, intermediate and high levels are triangles, circles and squares, respectively), which is evidence that either the receptor cluster size or the interaction strength (or both) increase with covalent modfication.
The most recent evidence suggests that the two predominant signaling states ('kinase-stimulated' and 'kinase-unstimulated') differ in density in the the membrane.
2. Bacterial Chemotaxis and Chemotaxis-like Signaling Pathways. Chemotaxis signaling pathways -- first and most thoroughly studied in E. coli and Salmonella, are widely distributed in prokaryotes; moreover many prokaryotic genomes have several chemotaxis gene clusters (see below). Pathways studied in other bacteria, for example Pseudomonas aeruginosa and Myxococcus xanthus, demonstrate that many 'chemotaxis-like' clusters -- perhaps the majority, regulate phenomena other than chemotaxis. In collaboration with Derek Lovley (Microbiology), we are studying the chemotaxis-like pathways of Geobacter; this genus of delta-proteobacteria are of interest for bioremediation and electricity generation in microbial fuel cells. The genomes of G. sulfurreducens, G. metallireducens and G. uraniireducens have several major chemotaxis gene clusters (Tran et al., in press), one to two clusters per genome resemble E. coli clusters (based on homology), one to two clusters resemble the frz and dif pathways of M. xanthus and P. aeruginosa, respectively. Based on homology and gene order, one cluster per genome looks to be unique to delta-protebacteria and still another unique to the Geobacter sp. The pathways unique to Geobacter are of particular interest, because these may have a unique role in bioremediation and electricity generation.
3. Functional Assembly of Membrane-Associated Proteins. The difficulties involved in isolation and study of transmembrane proteins complexes (left) has led us to a develop methods where the soluble portion of the membrane protein (the cytoplasmic domain of the aspartate receptor)
4. Membrane-Protein Interactions. In a joint project with the Dinsmore Group in the Physics Department, we seek to understand quantitatively and mechanistically, the energetics of membrane-protein interactions, which are manifested as changes in the phase separation properties and/or membrane curvature. The BAR superfamily are particularly interesting in this respect because of the role these proteins have in sensing and modulating membrane curvature. The images below, show a time series that records the transformation of a Giant Unilamellar Vesicle after the addition of amphiphysin BAR domain to the vesicle solution. (Images courtesy of Jaime Hutchison, Dinsmore Group, UMass Physics Department)