Robert K. Nakamoto
Primary AppointmentProfessor, Molecular Physiology and Biological Physics
- PhD, University of Maryland
- AB, Zoology, University of California, Berkeley, CA
- PhD, Biochemistry, University of Maryland, Baltimore, MD
- Postdoc, Biochemistry, Stanford University, Stanford, CA
- Postdoc, Biochemistry and Genetics, Yale University, New Haven, CT
Structure-Function of Active Transporters
Our goal is to understand the molecular mechanisms of these different transporters. We use a variety of biochemical, biophysical and structural approaches combined with genetic and molecular biological approaches to probe the structure-function relationships. In such ways, we can obtain measurements of the structural dynamics that occur during the transport cycle. The dynamics are correlated to the kinetics of the partial reactions occurring during transport and the energetics of these transitions. The data are used to generate models which can then be computationally simulated. These approaches allow us to understand how the transporters use the energy derived from chemical reactions or from electrochemical gradients to couple to the mechanical movement of molecules from one side of a membrane to the other. With high resolution structural data as a guide, we use site-directed mutagenesis to test our mechanistic models by altering single amino acids, or segments of the protein that carry out specific roles.
Not surprisingly, each of the transporters use vastly different molecular mechanisms. The P-glycoprotein binds substrate drugs from within the lipid bilayer and uses energy to rehydrate the transported compound on the exterior half of the membrane; the FOF1 transport and catalytic mechanisms are rotary motors which are coupled by a long coiled-coil structure akin to a drive shaft; and the BtuB outer membrane transporter interacts in a specific manner with the inner membrane protein TonB to activate the translocation of the large cyano-cobalamin molecule into the periplasmic space. In each case, the transporter mechanism is optimized for its specific physiological role.