The general research aims of my group are to use molecular modeling and bioinformatics to analyze structure, function, and molecular evolution of membrane proteins. Membrane proteins are one of the most important classes of proteins. They comprise about 30% of most genomes and are involved in many biological processes. They are especially important in biomedical research because most targets of current pharmaceutical projects are membrane proteins. Unfortunately, membrane protein structures are difficult to determine experimentally, and most that are determined come from prokaryotes. We fill some of this structural void by developing computational methods of analyzing sequences and developing structural models of membrane proteins. We use computational analyses to do the following:1)Address questions that are not answered by crystal structures. 2)Assist in understanding similarities and differences among homologous proteins.3)Relate structural and sequence information to functional properties.4)Assist in the design and interpretation of experimental studies.Our current projects involve developing models of the structure and gating mechanisms of potassium (K+) channels and their relatives.Potassium channels and related protein comprise the third largest superfamily of human genes. These proteins are found in almost all cells from bacteria on up. This category of membrane proteins contains several diverse superfamilies of channels including Na+, Ca2+, cyclic nucleotide-gated, TRP and its homologs, glutamate-activated, and Ca2+ release channels plus some K+ symporters and transporters. The smallest of these proteins are 2TM K+ channels that have four identical subunits; each of which has only two transmembrane helices, M1 and M2. A 'P' hairpin segment that spans only the outer half of the transmembrane region is located between M1 and M2. The P segment determines the selectivity of the channel. 6TM K+ channels are more complex, with each alpha subunit having four additional transmembrane segments, S1-S4, that precede the pore-forming S5-P-S6 motif (analogous to the M1-P-M2 motif of 2TM channels) and that forms a voltage-sensing domain in voltage-gated channels. Voltage-gated Ca2+ and Na+ channels have only one alpha subunit; however, it contains four homologous 6TM motifs. One of our goals is to develop structural models of the transmembrane region of at least one member of each major family of K+ channel related proteins. A major goal in membrane biophysics for over half a century has been to understand the molecular mechanisms by which voltage-dependent channels gate. We are developing three-dimensional models of these mechanisms. The importance of understanding the structure and functional mechanisms of K+ channels has been recognized by the awarding of the Nobel Prize in Chemistry to Roderick MacKinnon for the work in his laboratory in solving the crystal structures of K+ channels. We are utilizing their crystallographic data to develop structural models of the gating mechanisms of the crystallized KvAP and Kv1.2 channel proteins. We are also developing models of some of their eukaryotic homologs that have been studied extensively and that are important drug targets. The KvAP crystal structure presents a particularly interesting molecular modeling challenge. It is difficult to reconcile the crystal structure of the complete KvAP channel protein, and the paddle model of the voltage-dependent gating that MacKinnon's group developed based on this structure, with many experimental results and with basic principles of membrane protein energetics. We suspect that the voltage-sensing domain (S1-S4) of this structure is grossly distorted, but that a second crystal structure of an isolated voltage-sensing domain has a native open conformation.
