STRUCTURE/FUNCTION STUDY OF A PROSTAGLANDIN TRANSPORTER

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Dr. Brenda Chan received her M.D. from the Mount Sinai School of Medicine, was an Internal Medicine resident at Montefiore Medical Center, and has been a Nephrology Fellow at the Albert Einstein College of Medicine since 1993. For the past two years she has devoted greater than 80% of her time, in the sponsor's lab, to basic research on the molecular mechanisms of a novel prostaglandin (PG) transporter ("PGT"). Dr. Chan's immediate career goals are to devote at least 75% of her time over the next five years to bench research so as to foster development of her technical and conceptual skills in membrane transport and molecular biology. An intense training program will include 1) formal evaluation by an Advisory Committee; 2) involvement in the broad Einstein scientific community, including the group in membrane transport; 3) close supervision in the Sponsor's lab; 4) protection of 75% of her time for research. Dr. Chan's long-term career goals are to establish herself as an independent investigator in the area of renal organic anion membrane transport within an academic nephrology division. The Sponsor's lab studies two related organic anion transporters. "PGT", a PG transporter expressed strongly in the renal papilla, transports PGE2, PGF2alpha, and thromboxane B2, and appears to mediate PG metabolic clearance. "OATP", expressed in the S3 brush border, transports primarily conjugated steroids. Dr. Chan has characterized several mechanisms of PGT transport using the Xenopus oocyte expression system. More recently she has used site-directed mutagenesis to begin to address PGT's topology, binding site, and charged residues. The applicant's broad aims are to define the structure-function relationships of PGT so as to understand mechanisms of PG transport in particular, and of organic anion transport in general. The Specific aims of this proposal are: 1) To identify the membrane topology of PGT by epitope tagging and N-glycosylation scanning mutagenesis; 2) To characterize the substrate binding site and translocation pathway using cysteine mutagenesis and amino acid modifying reagents. The results will likely have broad and important implications for nephrology, and will allow a better general understanding of organic anion transport.