RESEARCH AREAS

Current Projects

1. Protein Binding & Structure-based Drug Design

Structural and Functional Mechanism of VR1. VR1 is one of the cloned nociceptive ion channels. It is highly expressed in sensory neurons and respond to both physical and chemical noxious stimuli, including heat, acidic pH, and irritant vanilloids (e.g. capsaicin, the pungent ingredient of hot peppers). Its diverse responsiveness has led to the suggestion that the receptor may act as a detector and integrator of noxious stimuli. Study of the receptor therefore provides an example to elucidate how nociception occurs at the first place. Our recent work focuses on the mechanisms of channel gating. Our goal is to establish, in molecular and mechanistic terms, how the channel is activated, how the physical and chemical activation pathways converge, and how the detection threshold is mediated by hyperalgesic factors (e.g. protons). Our approach involves patch-clamp recordings from recombinant channels in heterologous expression systems, combined with kinetic analysis to unravel the molecular events occurring during activation, along with mutagenesis to identify functional domains of the receptor.

2. Hidden Markov Modeling of Molecular Kinetics

Proteins respond to external factors through conformational changes. Static images as obtained by X-ray, NMR and microscopy provide a framework for understanding, but the dynamics that often determine functionality are rarely observed directly and can only be inferred from kinetic analysis of functional outputs. To this end, single molecule experiments provide unprecedented resolutions. Recording of currents from single ion channels has the most extensive history of single molecule techniques, but rapid progress has been made in recent years with other molecules using laser traps, single molecule fluorescence and atomic force microscopy. The measurements from such experiments contain a richness of kinetic detail about molecular structure and function that is difficult to obtain by other means.

Establishing molecular kinetics from single molecule activity, however, is a complicated process. Because molecules are embedded in a thermal bath, their activity is stochastic so that statistical tools are necessary for analysis. Further complicating the analysis are problems arising from limitations of instruments. Conformations that are kinetically different may give rise to the same functional observations, making transitions among them indistinguishable. The time scale of events in proteins is broad ranging from fs to seconds. The recording apparatus, however, has limited resolutions, which causes rapid transitions to go undetected. The noise in the recordings can be substantial, further contaminating the already small single molecule activity. We have been working on the problem for several years, with an emphasis on ion channel proteins. Our goal is to develop sophisticated analytical techniques that can take account of practical limitations while provide reliable estimates of kinetic properties. Some of the techniques that we have developed are already widely used by scientists around the world for studies of a variety of ion channels.