Coupling crystallography with kinetic studies leads to identification of a previously unknown binding pocket

Institute for Molecular Bioscience, The University of Queensland* National School of Pharmacy, University of Otago** Department of Medicinal Chemistry, The University of Kansas*** College of Pharmacy, University of Michigan****
â—‹Nyssa Drinkwater* Christine L Gee* Joel D Tyndall** Gary L Grunewald*** Wu Qian**** Michael J McLeish**** Jennifer L Martin*

Phenylethanolamine N-Methyltransferase (PNMT), as the enzyme that synthesises adrenaline in the central nervous system has been the target of structure-based inhibitor design for 25 years. Recently, on the basis of computational studies, inhibitors predicted to be too large to occupy the PNMT active site surprised researchers by maintaining high levels of potency. X-ray crystallographic structures of the enzyme:inhibitor complexes now show the reasons for this include dramatic movements of key active site residues and rigid body movements of nearby alpha helices. The described conformational changes reveal a previously unknown binding cavity, and result in a doubling in volume of the ligand-binding pocket. Furthermore, kinetic and mutational analysis shows that these major structural changes occur at relatively minor energetic cost. Presented here is a detailed structural, mutational, and kinetic study characterising the described structural perturbations. This research now has implications towards emerging techniques of drug design and development including virtual screening of compounds and fragment-based lead discovery.