Biomedical subjects

Philip A Gurnev

Publications and source records attributed to Philip A Gurnev.

6 recordsLinked to original sources

PubMed2006

Docking of a single phage lambda to its membrane receptor maltoporin as a time-resolved event.

We have been able to observe the first step in bacteriophage infection, the docking of phage lambda to its membrane receptor maltoporin, at the single-particle level. High-resolution conductance recording from a single trimeric maltoporin channel reconstituted into a planar lipid bilayer has allowed detection of the simultaneous and irreversible interaction of the phage tail with all three monomers of the receptor. The formation of a phage-maltoporin complex affects the channel transport properties. Our analysis demonstrates that phage attaches symmetrically to all three receptor monomers. The statistics of sugar binding to the phage-receptor complex on the side opposite to phage docking show that the monomers of maltoporin still bind sugar independently, with the kinetic constants expected from those of the phage-free receptor. This finding suggests that phage docking does not distort the structure of the receptor, and that the phage-binding regions are close to, but do not overlap with, the sugar-binding domains of the maltoporin monomers. However, ion fluxes through the pores of maltoporin in the phage-receptor complex share a new common pathway. We expect that the present study contributes to the current needs for structural information on the functional complexes involved in intercellular recognition.

Bacterial Outer Membrane Proteins↗

PubMed2006

Actin and amphiphilic polymers influence on channel formation by Syringomycin E in lipid bilayers.

The bacterial lipodepsipeptide syringomycin E (SRE) added to one (cis-) side of bilayer lipid membrane forms voltage dependent ion channels. It was found that G-actin increased the SRE-induced membrane conductance due to formation of additional SRE-channels only in the case when actin and SRE were applied to opposite sides of a lipid bilayer. The time course of conductance relaxation depended on the sequence of SRE and actin addition, suggesting that actin binds to the lipid bilayer and binding is a limiting step for SRE-channel formation. G-actin adsorption on the membrane was irreversible. The amphiphilic polymers, Konig's polyanion (KP) and poly(Lys, Trp) (PLT) produced the actin-like effect. It was shown that the increase in the SRE membrane activity was due to hydrophobic interactions between the adsorbing molecules and membrane. Nevertheless, hydrophobic interactions were not sufficient for the increase of SRE channel-forming activity. The dependence of the number of SRE-channels on the concentration of adsorbing species gave an S-shaped curve indicating cooperative adsorption of the species. Kinetic analysis of SRE-channel number growth led to the conclusion that the actin, KP, and PLT molecules form aggregates (domains) on the trans-monolayer. It is suggested that an excess of SRE-channel formation occurs within the regions of the cis-monolayer adjacent to the domains of the adsorbed molecules, which increase the effective concentration of SRE-channel precursors.

Actins↗

PubMed2006

Functional subconformations in protein folding: evidence from single-channel experiments.

We study fluctuations in ion conductance and enzymatic rates of the sugar-specific channel-forming membrane protein, Maltoporin, at the single-molecule level. Specifically, we analyze time-persistent deviations in the transport parameters of individual channels from the multichannel averages and discuss our findings in the context of static disorder in protein folding. We show that the disorder responsible for variations in ion conductance does not affect sugar binding, suggesting that Maltoporin can exist in a wide set of fully functional, yet distinctly different, subconformations.

Bacterial Outer Membrane Proteins↗

PubMed2003

Effective gating charge of ion channels induced by toxin syringomycin E in lipid bilayers.

To elucidate the voltage gating of syringomycin E (SRE) ion channels in lipid bilayers, the effective gating charge q was measured under different conditions. It was shown that q and its sign are dependent on membrane surface charge, dipole potential, and the outer potential (Delta phi). The q values were positive for charged bilayers and negative for uncharged bilayers bathed in the same 0.1 M NaCl solutions. Effects of dipole modifying agents on the gating properties of SRE channels were measured. In uncharged bilayers, addition of phloretin resulted in an increase of q values. For charged bilayers, the presence of RH-421 or 6-ketocholestanol leads to the reverse in the sign of q from positive to negative. The q values were potential-dependent at higher negative voltages with charged membranes bathed in solutions with high salt concentrations. It is concluded that lipid molecules participating in the SRE channel structure contribute to channel formation work due to Coulomb and dipolar interactions with the electric field applied to a membrane. The potential dependence of q is explained by interactions of charged and uncharged lipids with SRE molecules in the channels.

Ion Channel Gating↗

PubMed2002

Syringotoxin pore formation and inactivation in human red blood cell and model bilayer lipid membranes.

The effect of syringotoxin (ST), a member of the cyclic lipodepsipeptides family (CLPs) produced by Pseudomonas syringae pv. syringae on the membrane permeability of human red blood cells (RBCs) and model bilayer lipid membranes (BLMs) was studied and compared to that of two recently investigated CLPs, syringomycin E (SRE) and syringopeptin 22A (SP22A) [Biochim. Biophys. Acta 1466 (2000) 79 and Bioelectrochemistry 52 (2000) 161]. The permeability-increasing effect of ST on RBCs was the least among the three CLPs. A time-dependent ST pore inactivation was observed on RBCs at 20 and 37 degrees C but not at 8 degrees C. From the kinetic model worked out parameters as permeability coefficient of RBC membrane for 86Rb(+) and pores mean lifetime were calculated. A shorter pores mean lifetime was calculated at 37 degrees C then at 20 degrees C, which gave us an explanation for the unusual slower rate of tracer efflux measured at 37 degrees C then that at 20 degrees C. The results obtained on BLM showed that the pore inactivation was due to a decrease in the number of pores but not to a change of their dwell time or conductance.

Cell Membrane Permeability↗

PubMed2002

Syringomycin E channel: a lipidic pore stabilized by lipopeptide?

Highly reproducible ion channels of the lipopeptide antibiotic syringomycin E demonstrate unprecedented involvement of the host bilayer lipids. We find that in addition to a pronounced influence of lipid species on the open-channel ionic conductance, the membrane lipids play a crucial role in channel gating. The effective gating charge, which characterizes sensitivity of the conformational equilibrium of the syringomycin E channels to the transmembrane voltage, is modified by the lipid charge and lipid dipolar moment. We show that the type of host lipid determines not only the absolute value but also the sign of the gating charge. With negatively charged bilayers, the gating charge sign inverts with increased salt concentration or decreased pH. We also demonstrate that the replacement of lamellar lipid by nonlamellar with the negative spontaneous curvature inhibits channel formation. These observations suggest that the asymmetric channel directly incorporates lipids. The charges and dipoles resulting from the structural inclusion of lipids are important determinants of the overall energetics that underlies channel gating. We conclude that the syringomycin E channel may serve as a biophysical model to link studies of ion channels with those of lipidic pores in membrane fusion.

Anti-Bacterial Agents↗