OVER-THE-BARRIER TRANSITION STATE ANALOGUES AND CRYSTAL STRUCTURE WITH MYCOBACTERIUM TUBERCULOSIS PURINE NUCLEOSIDE PHOSPHORYLASE

摘要: Stable chemical analogues of enzymatic transition states are imperfect mimics since they lack the partial bond character state. We synthesized structural variants Immucillins as state for purine nucleoside phosphorylase and characterized them with enzyme from Mycobacterium tuberculosis (MtPNP). PNPs form ribooxacarbenium ion catalyze nucleophilic displacement reactions by migration cationic carbon between enzymatically immobilized phosphate nucleophiles. As bond-breaking progresses, carbocation builds on ribosyl group, distance increases, anion decreases. Transition were produced increased incorporating a methylene bridge these groups. Immucillin-H (ImmH), DADMe-ImmH, DADMe-ImmG mimic MtPNP slow-onset, tight-binding inhibitors equilibrium dissociation constants 650, 42, 24 pM. Crystal structures complexes ImmH DADMe-ImmH reveal an ion-pair inhibitor cation phosphoryl anion. The stronger (2.7 A) is found DADMe-ImmH. position bound resembles substrate side barrier, more closely product barrier. ability to probe both sides barrier provides expanded opportunities explore analogue design in N-ribosyltransferases. This approach has resulted highest affinity known MtPNP.