substrate binding mode and its implication on drug design for botulinum neurotoxin a衬底绑定模式及其对药物设计的含义为肉毒神经毒素.pdfVIP

Substrate Binding Mode and Its Implication on Drug Design for Botulinum Neurotoxin A 1 1 2 1 Desigan Kumaran , Richa Rawat , S. Ashraf Ahmed , Subramanyam Swaminathan * 1 Biology Department, Brookhaven National Laboratory, Upton, New York, United States of America, 2 Integrated Toxicology Division, US Army Research Institute of Infectious Diseases, Fort Detrick, Maryland, United States of America Abstract The seven antigenically distinct serotypes of Clostridium botulinum neurotoxins, the causative agents of botulism, block the neurotransmitter release by specifically cleaving one of the three SNARE proteins and induce flaccid paralysis. The Centers for Disease Control and Prevention (CDC) has declared them as Category A biowarfare agents. The most potent among them, botulinum neurotoxin type A (BoNT/A), cleaves its substrate synaptosome-associated protein of 25 kDa (SNAP-25). An efficient drug for botulism can be developed only with the knowledge of interactions between the substrate and enzyme at the active site. Here, we report the crystal structures of the catalytic domain of BoNT/A with its uncleavable SNAP-25 peptide 197 202 197 202 ˚ ˚ QRATKM and its variant RRATKM to 1.5 A and 1.6 A, respectively. This is the first time the structure of an uncleavable substrate bound to an active botulinum neurotoxin is reported and it has helped in unequivocally defining S1 to S59 sites. These substrate peptides make interactions with the enzyme predominantly by the residues from 160, 200, 250 and 370 loops. Most notably, the amino nitrogen and carbonyl oxygen of P1 residue (Gln197) chelate the zinc ion and replace the