1N3I

Crystal Structure of Mycobacterium tuberculosis PNP with transition state analog DADMe-ImmH

Summary for 1N3I

• Entry DOI: 10.2210/pdb1n3i/pdb
• Related: 1G2O
• Descriptor: Purine Nucleoside Phosphorylase, PHOSPHATE ION, 7-[[(3R,4R)-3-(hydroxymethyl)-4-oxidanyl-pyrrolidin-1-ium-1-yl]methyl]-3,5-dihydropyrrolo[3,2-d]pyrimidin-4-one, ... (4 entities in total)
• Functional Keywords: transition state complex, trimer, pnp, transferase
• Biological source: Mycobacterium tuberculosis
• Total number of polymer chains: 3
• Total formula weight: 83879.15

Authors

Lewandowicz, A.,Shi, W.,Evans, G.B.,Tyler, P.C.,Furneaux, R.H.,Basso, L.A.,Santos, D.S.,Almo, S.C.,Schramm, V.L. (deposition date: 2002-10-28, release date: 2003-09-30, Last modification date: 2023-10-25)

Primary citation

Lewandowicz, A.,Shi, W.,Evans, G.B.,Tyler, P.C.,Furneaux, R.H.,Basso, L.A.,Santos, D.S.,Almo, S.C.,Schramm, V.L.
Over-The-Barrier Transition State Analogues Provide New Chemistries for Inhibitor Design: The Case of Purine Nucleoside Phosphorylase
BIOCHEMISTRY, 42:6057-6066, 2003

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

PubMed: 12755607
DOI: 10.1021/bi0343830

Experimental method

X-RAY DIFFRACTION (1.9 Å)