Biomedical subjects

Robert G Endres

Publications and source records attributed to Robert G Endres.

6 recordsLinked to original sources

PubMed2006

Kinetic analysis of the assembly of the outer membrane protein LamB in Escherichia coli mutants each lacking a secretion or targeting factor in a different cellular compartment.

Outer membrane beta-barrel proteins in gram-negative bacteria, such as Escherichia coli, must be translocated from their site of synthesis in the cytoplasm to the periplasm and finally delivered to the outer membrane. At least a dozen proteins located in the cytoplasm, the periplasm, and both the inner and outer membranes are required to catalyze this complex assembly process. At normal growth temperatures and conditions the transport and assembly processes are so fast that assembly intermediates cannot be detected. Using cells grown at a low temperature to slow the assembly process and pulse-chase analysis with immunodetection methods, we followed newly synthesized LamB molecules during their transit through the cell envelope. The quality and reproducibility of the data allowed us to calculate rate constants for three different subassembly reactions. This kinetic analysis revealed that secB and secD mutants exhibit nearly identical defects in precursor translocation from the cytoplasm. However, subsequent subassembly reaction rates provided no clear evidence for an additional role for SecD in LamB assembly. Moreover, we found that surA mutants are qualitatively indistinguishable from yfgL mutants, suggesting that the products of both of these genes share a common function in the assembly process, most likely the delivery of LamB to the YaeT assembly complex in the outer membrane.

Bacterial Outer Membrane Proteins↗

PubMed2006

Precise adaptation in bacterial chemotaxis through "assistance neighborhoods".

The chemotaxis network in Escherichia coli is remarkable for its sensitivity to small relative changes in the concentrations of multiple chemical signals over a broad range of ambient concentrations. Key to this sensitivity is an adaptation system that relies on methylation and demethylation (or deamidation) of specific modification sites of the chemoreceptors by the enzymes CheR and CheB, respectively. It was recently discovered that these enzymes can access five to seven receptors when tethered to a particular receptor. We show that these "assistance neighborhoods" are necessary for precise adaptation in a model for signaling by clusters of chemoreceptors. In agreement with experiment, model clusters composed of receptors of different types exhibit high sensitivity and precise adaptation over a wide range of chemical concentrations and the response of adapted clusters to addition/removal of attractant scales with free-energy change. We predict two limits of precise adaptation at large attractant concentrations: Either receptors reach full methylation and turn off, or receptors become saturated and cease to respond to attractant but retain their adapted activity.

Adaptation, Physiological↗

PubMed2006

Weight matrices for protein-DNA binding sites from a single co-crystal structure.

Transcription-factor proteins bind to specific DNA sequences to regulate gene expression in cells. DNA-binding sites are often identified using weight matrices calculated from multiple known binding sites. However, in many cases the number of examples is limited. Here, we report on an atomistic method that starts from an x-ray co-crystal structure of the protein bound to one particular DNA sequence, and infers other binding sites, which are used to construct a weight matrix. The emphasis of the paper is on using the Wang-Landau Monte Carlo algorithm to efficiently sample high-affinity binding sites, which demonstrates that sampling can produce accurate weight matrices in analogy to bioinformatics approaches. For cases of low complexity, we compare to the exhaustive (but slow) dead-end elimination algorithm. To recover crystal binding sites, it is important to include bound water in the protein-DNA interface. Our approach can, in principle, even be applied when no native protein-DNA co-crystal structure is available, only the structure of a closely related homologous protein whose amino-acid sequence is changed to the protein of interest.

Algorithms↗

PubMed2006

Receptor-receptor coupling in bacterial chemotaxis: evidence for strongly coupled clusters.

Receptor coupling is believed to explain the high sensitivity of the Escherichia coli chemotaxis network to small changes in levels of chemoattractant. We compare in detail the activity response of coupled two-state receptors for different models of receptor coupling: weakly-coupled extended one-dimensional and two-dimensional lattice models and the Monod-Wyman-Changeux model of isolated strongly-coupled clusters. We identify features in recent data that distinguish between the models. Specifically, researchers have measured the receptor activity response to steps of chemoattractant for a variety of engineered E. coli strains using in vivo fluorescence resonance energy transfer. We find that the fluorescence resonance energy transfer results for wild-type and for a low-activity mutant are inconsistent with the lattice models of receptor coupling, but consistent with the Monod-Wyman-Changeux model of receptor coupling, suggesting that receptors form isolated strongly-coupled clusters.

Binding Sites↗

PubMed2006

Chemosensing in Escherichia coli: two regimes of two-state receptors.

The chemotaxis network in Escherichia coli is remarkable for its sensitivity to small relative changes in the concentrations of multiple chemical signals. We present a model for signal integration by mixed clusters of interacting two-state chemoreceptors. Our model results compare favorably to the results obtained by Sourjik and Berg with in vivo fluorescence resonance energy transfer. Importantly, we identify two distinct regimes of behavior, depending on the relative energies of the two states of the receptors. In regime I, coupling of receptors leads to high sensitivity, while in regime II, coupling of receptors leads to high cooperativity, i.e., high Hill coefficient. For homogeneous receptors, we predict an observable transition between regime I and regime II with increasing receptor methylation or amidation.

Chemotaxis↗

PubMed2004

Toward an atomistic model for predicting transcription-factor binding sites.

Identifying the specific DNA-binding sites of transcription-factor proteins is essential to understanding the regulation of gene expression in the cell. Bioinformatics approaches are fast compared to experiments, but require prior knowledge of multiple binding sites for each protein. Here, we present an atomistic force-field method to predict binding sites based only on the X-ray structure of a related bound complex. Specific flexible contacts between the protein and DNA are modeled by a library of amino acid side-chain rotamers. Using the example of the mouse transcription factor, Zif268, a well-studied zinc-finger protein, we show that the protein sequence alone, without the detailed experimental structure, gives a strong bias toward the consensus binding site.

Algorithms↗