COMPARISON OF THE CONSTRUCTION OF A 3-D MODEL FOR HUMAN THROMBOXANE SYNTHASE USING P450CAM AND BM-3 AS TEMPLATES - IMPLICATIONS FOR THE SUBSTRATE-BINDING POCKET

A 3-D model of human thromboxane A(2) synthase (TXAs) was constructed using a homology modeling approach based on information from the 2.0 Angstrom crystal structure of the hemoprotein domains of cytochrome P450BM-3 and P450cam. P450BM-3 is a bacterial fatty acid monooxygenase resembling eukaryotic microsomal cytochrome P450s in primary structure and function. TXAS shares 26.4% residue identity and 48.4% residue similarity with the P450BM-3 hemoprotein domain, The homology score between TXAS and P450BM-3 is much higher than that between TXAS and P450cam. Alignment between TXAS and the P450BM-3 hemoprotein domain or P450cam was determined through sequence searches. The P450BM-3 or P450cam main-chain coordinates were applied to the TXAS main chain in those segments where the two sequences were well aligned. These segments were linked to one another using a fragment search method, and the side chains were added to produce a 3-D model for TXAS. A TXAS substrate, prostaglandin H-2 (PGH(2)) was docked into the TXAS cavity corresponding to the arachidonic acid binding pocket in P450BM-3 or camphor binding site in P450cam. Regions of the heme and putative PGH(2) binding cavities in the TXAS model were identified and analyzed. The segments and residues involved in the active-site pocket of the TXAS model provide reasonable candidates for TXAS protein engineering and inhibitor design. Comparison of the TXAS model based on P450BPM-3 with another TXAS model based on the P450cam structure indicated that P450BM-3 is a more suitable template for homology modeling of TXAS.