STRUCTURAL STUDIES ON AN INHIBITORY ANTIBODY AGAINST THERMUS-AQUATICUS DNA-POLYMERASE SUGGEST MODE OF INHIBITION

TP7, an antibody against Thermus aquaticus DNA polymerase I (TaqP), is used as a thermolabile switch in 'hot start' variations of PCR to min imize non-specific amplification events. Earlier studies have establis hed that TP7 binds to the polymerase domain of TaqP, competes with pri mer template complex for binding and is a potent inhibitor of the poly merase activity of TaqP, We report crystallographic determination of t he structure of an Fab fragment of TP7 and computational docking of th e structure with the known three-dimensional structure of the enzyme. Our observations strongly suggest that the origin of inhibitory abilit y of TP7 is its binding to enzyme residues involved in DNA binding and polymerization mechanism. Although criteria unbiased by extant bioche mical data have been used in identification of a putative solution, th e resulting complex offers an eminently plausible structural explanati on of biochemical observations. The results presented are of general s ignificance for interpretation of docking experiments and in design of small molecular inhibitors of TaqP, that are not structurally similar to substrates, for use in PCR, structural and functional similarities noted among DNA polymerases, and the fact that several DNA polymerase s are pharmacological targets, make discovery of non-substrate based i nhibitors of additional importance.