Several diseases, including cystic fibrosis, immunodeficiency, and multidrug-resistant cancer, are linked to a family of ubiquitous membrane proteins called ABC (ATP-binding cassette) transporters. ABC transporters are multi-domain proteins that pump a variety of substrates across biological membranes. This proposal is designed to elucidate the molecular mechanism underlying the active transport process mediated by ABC transporters, using the E. coli maltose transporter as a model system. Active transport appears to occur through an alternating access mechanism driven by ATP hydrolysis, in which the transporter cycles between two conformations, each exposing an internal substrate-binding site to only one side of the membrane. Biochemistry, electron paramagnetic resonance and X-ray crystallography experiments are designed to elucidate details of this process with a major focus on the identification of new structural intermediates in the transport cycle, including those that may be important in the initiation of the transport cycle (Aim 1) and those that may lie further along the pathway between the transition and resting state (Aim 2). In addition, we will build on knowledge gained from crystal structures to investigate details about the coupling of ATP hydrolysis to translocation (Aim 3) and how different substrates are recognized by the transporter (Aim 4). Because all members of the ABC transporter superfamily share sequence similarity and a common domain organization, these experiments will advance our general understanding of how conformational changes of ABC transporters result in substrate translocation. Research emphasizing basic mechanisms, as revealed by a combination of biochemistry and high-resolution structures, will accelerate the design of drug therapies and interventions to improve the health of individuals who suffer from the myriad of diseases associated with ABC transporters.
ATP-binding cassette (ABC) transporters are membrane proteins that selectively transport substrates across biological membranes. Many are involved in disease, including drug-resistant cancer, cystic fibrosis, diabetes, macular degeneration and atherosclerosis. Research emphasizing basic mechanisms, as revealed by a combination of biochemical and high-resolution structures, will accelerate optimal design of drug therapies and interventions to improve human health.
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