One-third-the-sites transition-state inhibitors for purine nucleoside phosphorylase

Robert W. Miles, Peter C. Tyler, Richard H. Furneaux, Carey K. Bagdassarian, Vern L. Schramm

Research output: Contribution to journalArticle

237 Citations (Scopus)

Abstract

Genetic defects in human purine nucleoside phosphorylase cause T-cell deficiency as the major phenotype. It has been proposed that efficient inhibitors of the enzyme might intervene in disorders of T-cell function. Compounds with features of the transition-state structure of purine nucleoside phosphorylase were synthesized and tested as inhibitors. The transition-state structure for purine nucleoside phosphorylase is characterized by (1) an elevated pK(a) at N7 of the purine ring for protonation or favorable H-bond interaction with the enzyme and (2) oxocarbenium ion formation in the ribosyl ring (Kline, P. C., and Schramm, V. L. (1995) Biochemistry 34, 1153-1162). Both features have been incorporated into the stable transition-state analogues, (1S)-1-(9-deazahypoxanthin-9- yl)-1,4-dideoxy-1,4-imino-D-ribitol (immucillin-H) and (1S)-1-(9-deazaguanin- 9-yl)-1,4-dideoxy-1,4-imino-D-ribitol (immucillin-G). Both inhibitors exhibit slow-onset tight-binding inhibition of calf spleen and human erythrocyte purine nucleoside phosphorylase. The inhibitors exhibit equilibrium dissociation constants (K(i)*) from 23 to 72 pM and are the most powerful inhibitors reported for the enzyme. Complete inhibition of the homotrimeric enzyme occurs at one mole of inhibitor per mole of enzymic trimer. Binding of the transition-state inhibitor at one site per trimer prevents inhibitor binding at the remaining two sites of the homotrimer. A mechanism of sequential catalysis at each subunit, similar to that of F1 ATPase, is supported by these results. Slow inhibitor dissociation (e.g., t 1/4 of 4.8 h) suggests that these compounds will have favorable pharmacologic properties. Interaction of transition-state inhibitors with purine nucleoside phosphorylase is different from reactant-state (substrate and product analogue) inhibitors of the enzyme which bind equally to all subunits of the homotrimer.

Original languageEnglish (US)
Pages (from-to)8615-8621
Number of pages7
JournalBiochemistry
Volume37
Issue number24
DOIs
StatePublished - Jun 16 1998

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Purine-Nucleoside Phosphorylase
Enzyme Inhibitors
Ribitol
T-cells
T-Lymphocytes
Biochemistry
Proton-Translocating ATPases
Protonation
Enzymes
Catalysis
Spleen
Erythrocytes
Ions
Phenotype
Defects
Substrates

ASJC Scopus subject areas

  • Biochemistry

Cite this

One-third-the-sites transition-state inhibitors for purine nucleoside phosphorylase. / Miles, Robert W.; Tyler, Peter C.; Furneaux, Richard H.; Bagdassarian, Carey K.; Schramm, Vern L.

In: Biochemistry, Vol. 37, No. 24, 16.06.1998, p. 8615-8621.

Research output: Contribution to journalArticle

Miles, RW, Tyler, PC, Furneaux, RH, Bagdassarian, CK & Schramm, VL 1998, 'One-third-the-sites transition-state inhibitors for purine nucleoside phosphorylase', Biochemistry, vol. 37, no. 24, pp. 8615-8621. https://doi.org/10.1021/bi980658d
Miles, Robert W. ; Tyler, Peter C. ; Furneaux, Richard H. ; Bagdassarian, Carey K. ; Schramm, Vern L. / One-third-the-sites transition-state inhibitors for purine nucleoside phosphorylase. In: Biochemistry. 1998 ; Vol. 37, No. 24. pp. 8615-8621.
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