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Laboratory of Membrane Biology and Biophysics

Research in my lab centers on ATP-binding cassette (ABC) transporters, a diverse group of membrane proteins integral to almost every biological process. In prokaryotes, these proteins are critical for survival. In humans, ABC transporters make up one of the largest gene families, and more than a dozen genetic diseases have been traced to ABC transporter defects. ABC transporters are also central to multidrug resistance in many pathogenic bacteria and in tumor cells. By pursing structural and mechanistic studies of ABC transporters, we hope to understand how nature utilizes the chemical energy of ATP hydrolysis to perform work – transporting substrates against their chemical gradients.

There are some examples of our projects:

The E. coli maltose transporter, a prototype for ABC transporters

malFigure 1. The maltose transporter.  Click here to see a movie of the full transport cycle.

The maltose transporter is a nutrient importer that has been extensively studied for over 40 years as a model system for membrane transporters. My lab has previously determined the crystal structures of the maltose transporter in different stages of the transport cycle, including the inward-facing resting state; the substrate-induced pre-translocation state; the outward-facing catalytic intermediate; and the EIIAGlc-inhibited state. Based on these structures and the rich genetic and biochemical data in the literature, we are now able to describe the atomic details of how ATP hydrolysis enables maltose transport; the chemistry of ATP hydrolysis; how different substrates are recognized; and, finally, how bacteria regulate maltose uptake to optimize growth. The mechanistic insights gleaned from the maltose transporter can also be applied to many other ABC transporters.

The maltose transporter is a nutrient importer that has been extensively studied for over 40 years as a model system for membrane transporters. My lab has previously determined the crystal structures of the maltose transporter in different stages of the transport cycle, including the inward-facing resting state; the substrate-induced pre-translocation state; the outward-facing catalytic intermediate; and the EIIAGlc-inhibited state. Based on these structures and the rich genetic and biochemical data in the literature, we are now able to describe the atomic details of how ATP hydrolysis enables maltose transport; the chemistry of ATP hydrolysis; how different substrates are recognized; and, finally, how bacteria regulate maltose uptake to optimize growth. The mechanistic insights gleaned from the maltose transporter can also be applied to many other ABC transporters.

P-glycoprotein, the multidrug transporter

Cancer cells develop resistance to a large number of chemically diverse compounds, a phenomenon known as multidrug resistance (MDR). In 1976, Juliano and Ling identified a glycoprotein enriched in colchicine-resistant cells but not in wild-type cells. The protein was named "P-glycoprotein" (P stands for permeability) because it was thought to confer drug resistance by making the membrane less permeable. This was a reasonable conclusion at the time, as it is difficult to imagine how a single transporter could recognize many unrelated drugs. It is now known that P-glycoprotein is an ABC transporter whose physiological function is to protect sensitive tissues and the fetus from toxicity. Our lab has made a long-term commitment to detailed structure/function studies of this important protein. We hope that this research will advance our understanding of the fundamental mechanisms underlying multidrug resistance and ultimately lead to novel therapeutic reagents for cancer treatment.

sopaFigure 2. P-glycoprotein. (a) Ribbon presentation of the structure. (b) Drug-stimulated ATPase activities in isolated membranes (blue) and in detergents (green).

PCAT: ABC transporters that process and secrete proteins

Some of the most challenging questions in the membrane transport field concern polypeptides. How does the transport system pass through large proteins across the membrane without destroying the integrity of the cell? How are the vast variety of protein substrates recognized? And what are the driving forces that define the directionality of the transport process?

 

The simplest protein secretion system is PCAT, the peptidase-containing ABC transporter that processes and transports polypeptide substrates. In gram-positive bacteria, PCATs export quorum-sensing or antimicrobial peptides. In gram-negative bacteria, PCAT proteins interact with an accessory protein and the outer membrane TolC to assemble a continuous channel penetrating both the inner and outer membranes. Substrates secreted by these ABC transporters vary from small bacteriocins to large toxins of several thousand residues. Our aim is to correlate structural and functional studies of PCAT to build a complete understanding of its function.

 

Research in my lab centers on ATP-binding cassette (ABC) transporters, a diverse group of membrane proteins integral to almost every biological process. In prokaryotes, these proteins are critical for