Isozyme-specific transition state inhibitors for the trypanosomal nucleoside hydrolases

David W. Parkin, Gerrit Limberg, Peter C. Tyler, Richard H. Furneaux, Xiang Yang Chen, Vern L. Schramm

Research output: Contribution to journalArticle

45 Citations (Scopus)

Abstract

Protozoan parasites lack de novo purine biosynthesis and require purine salvage from the host. Nucleoside hydrolases are involved in nucleoside salvage and are not found in mammals, making them protozoan-specific targets for inhibitor design. Several protozoan nucleoside hydrolase isozymes with distinct substrate specificities have been characterized. Novel substituted iminoribitols have been synthesized to resemble the transition state structure of the nonspecific inosine-uridine nucleoside hydrolase from Crithidia fasciculata (IU-nucleoside hydrolase). These inhibitors have been characterized for this enzyme and for a purine-specific nucleoside hydrolase (IAG-nucleoside hydrolase) from Trypanosoma brucei brucei. Inhibitors which provide nanomolar inhibition constants for IU-nucleoside hydrolase exhibit micromolar inhibition constants for the IAG-enzyme. For example, p- bromophenyliminoribitol inhibits the IU- and IAG-enzymes with dissociation constants of 28 nM and 190 μM, respectively. Substrate specificity, the action of transition state inhibitors and the pH-dependence of the kinetic constants establish that the catalytic mechanisms and transition state structures are fundamentally different for the IU- and IAG-isozymes. The finding is remarkable since these isozymes share significant homology at the catalytic sites and both use inosine as a preferred substrate. The specificity of the transition state analogues indicates that logically- designed transition state inhibitors are isozyme-specific, with (K(m)/K(i) IU-nucleoside hydrolase)/(K(m)/K(i) IAG-nucleoside hydrolase) values up to 39 000. The mechanism of the differential inhibition is based on the relative leaving group activation and ribosyl-oxocarbenium-forming abilities of these enzymes. In addition to providing isozyme-specific inhibitors, the novel molecules described here have diagnostic value for the nature of the transition states for N-ribohydrolase enzymes.

Original languageEnglish (US)
Pages (from-to)3528-3534
Number of pages7
JournalBiochemistry
Volume36
Issue number12
DOIs
StatePublished - Mar 25 1997

Fingerprint

N-Glycosyl Hydrolases
Inosine
Crithidia fasciculata
Isoenzymes
Uridine
Enzymes
Salvaging
Substrate Specificity
Substrates
Hydrolases
Trypanosoma brucei brucei
Mammals
Biosynthesis
Nucleosides
Catalytic Domain
Parasites
Chemical activation
Molecules
Kinetics
purine

ASJC Scopus subject areas

  • Biochemistry

Cite this

Parkin, D. W., Limberg, G., Tyler, P. C., Furneaux, R. H., Chen, X. Y., & Schramm, V. L. (1997). Isozyme-specific transition state inhibitors for the trypanosomal nucleoside hydrolases. Biochemistry, 36(12), 3528-3534. https://doi.org/10.1021/bi962319v

Isozyme-specific transition state inhibitors for the trypanosomal nucleoside hydrolases. / Parkin, David W.; Limberg, Gerrit; Tyler, Peter C.; Furneaux, Richard H.; Chen, Xiang Yang; Schramm, Vern L.

In: Biochemistry, Vol. 36, No. 12, 25.03.1997, p. 3528-3534.

Research output: Contribution to journalArticle

Parkin, DW, Limberg, G, Tyler, PC, Furneaux, RH, Chen, XY & Schramm, VL 1997, 'Isozyme-specific transition state inhibitors for the trypanosomal nucleoside hydrolases', Biochemistry, vol. 36, no. 12, pp. 3528-3534. https://doi.org/10.1021/bi962319v
Parkin, David W. ; Limberg, Gerrit ; Tyler, Peter C. ; Furneaux, Richard H. ; Chen, Xiang Yang ; Schramm, Vern L. / Isozyme-specific transition state inhibitors for the trypanosomal nucleoside hydrolases. In: Biochemistry. 1997 ; Vol. 36, No. 12. pp. 3528-3534.
@article{83b4b58d4b0d41d7827f40e7340a2b4e,
title = "Isozyme-specific transition state inhibitors for the trypanosomal nucleoside hydrolases",
abstract = "Protozoan parasites lack de novo purine biosynthesis and require purine salvage from the host. Nucleoside hydrolases are involved in nucleoside salvage and are not found in mammals, making them protozoan-specific targets for inhibitor design. Several protozoan nucleoside hydrolase isozymes with distinct substrate specificities have been characterized. Novel substituted iminoribitols have been synthesized to resemble the transition state structure of the nonspecific inosine-uridine nucleoside hydrolase from Crithidia fasciculata (IU-nucleoside hydrolase). These inhibitors have been characterized for this enzyme and for a purine-specific nucleoside hydrolase (IAG-nucleoside hydrolase) from Trypanosoma brucei brucei. Inhibitors which provide nanomolar inhibition constants for IU-nucleoside hydrolase exhibit micromolar inhibition constants for the IAG-enzyme. For example, p- bromophenyliminoribitol inhibits the IU- and IAG-enzymes with dissociation constants of 28 nM and 190 μM, respectively. Substrate specificity, the action of transition state inhibitors and the pH-dependence of the kinetic constants establish that the catalytic mechanisms and transition state structures are fundamentally different for the IU- and IAG-isozymes. The finding is remarkable since these isozymes share significant homology at the catalytic sites and both use inosine as a preferred substrate. The specificity of the transition state analogues indicates that logically- designed transition state inhibitors are isozyme-specific, with (K(m)/K(i) IU-nucleoside hydrolase)/(K(m)/K(i) IAG-nucleoside hydrolase) values up to 39 000. The mechanism of the differential inhibition is based on the relative leaving group activation and ribosyl-oxocarbenium-forming abilities of these enzymes. In addition to providing isozyme-specific inhibitors, the novel molecules described here have diagnostic value for the nature of the transition states for N-ribohydrolase enzymes.",
author = "Parkin, {David W.} and Gerrit Limberg and Tyler, {Peter C.} and Furneaux, {Richard H.} and Chen, {Xiang Yang} and Schramm, {Vern L.}",
year = "1997",
month = "3",
day = "25",
doi = "10.1021/bi962319v",
language = "English (US)",
volume = "36",
pages = "3528--3534",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "12",

}

TY - JOUR

T1 - Isozyme-specific transition state inhibitors for the trypanosomal nucleoside hydrolases

AU - Parkin, David W.

AU - Limberg, Gerrit

AU - Tyler, Peter C.

AU - Furneaux, Richard H.

AU - Chen, Xiang Yang

AU - Schramm, Vern L.

PY - 1997/3/25

Y1 - 1997/3/25

N2 - Protozoan parasites lack de novo purine biosynthesis and require purine salvage from the host. Nucleoside hydrolases are involved in nucleoside salvage and are not found in mammals, making them protozoan-specific targets for inhibitor design. Several protozoan nucleoside hydrolase isozymes with distinct substrate specificities have been characterized. Novel substituted iminoribitols have been synthesized to resemble the transition state structure of the nonspecific inosine-uridine nucleoside hydrolase from Crithidia fasciculata (IU-nucleoside hydrolase). These inhibitors have been characterized for this enzyme and for a purine-specific nucleoside hydrolase (IAG-nucleoside hydrolase) from Trypanosoma brucei brucei. Inhibitors which provide nanomolar inhibition constants for IU-nucleoside hydrolase exhibit micromolar inhibition constants for the IAG-enzyme. For example, p- bromophenyliminoribitol inhibits the IU- and IAG-enzymes with dissociation constants of 28 nM and 190 μM, respectively. Substrate specificity, the action of transition state inhibitors and the pH-dependence of the kinetic constants establish that the catalytic mechanisms and transition state structures are fundamentally different for the IU- and IAG-isozymes. The finding is remarkable since these isozymes share significant homology at the catalytic sites and both use inosine as a preferred substrate. The specificity of the transition state analogues indicates that logically- designed transition state inhibitors are isozyme-specific, with (K(m)/K(i) IU-nucleoside hydrolase)/(K(m)/K(i) IAG-nucleoside hydrolase) values up to 39 000. The mechanism of the differential inhibition is based on the relative leaving group activation and ribosyl-oxocarbenium-forming abilities of these enzymes. In addition to providing isozyme-specific inhibitors, the novel molecules described here have diagnostic value for the nature of the transition states for N-ribohydrolase enzymes.

AB - Protozoan parasites lack de novo purine biosynthesis and require purine salvage from the host. Nucleoside hydrolases are involved in nucleoside salvage and are not found in mammals, making them protozoan-specific targets for inhibitor design. Several protozoan nucleoside hydrolase isozymes with distinct substrate specificities have been characterized. Novel substituted iminoribitols have been synthesized to resemble the transition state structure of the nonspecific inosine-uridine nucleoside hydrolase from Crithidia fasciculata (IU-nucleoside hydrolase). These inhibitors have been characterized for this enzyme and for a purine-specific nucleoside hydrolase (IAG-nucleoside hydrolase) from Trypanosoma brucei brucei. Inhibitors which provide nanomolar inhibition constants for IU-nucleoside hydrolase exhibit micromolar inhibition constants for the IAG-enzyme. For example, p- bromophenyliminoribitol inhibits the IU- and IAG-enzymes with dissociation constants of 28 nM and 190 μM, respectively. Substrate specificity, the action of transition state inhibitors and the pH-dependence of the kinetic constants establish that the catalytic mechanisms and transition state structures are fundamentally different for the IU- and IAG-isozymes. The finding is remarkable since these isozymes share significant homology at the catalytic sites and both use inosine as a preferred substrate. The specificity of the transition state analogues indicates that logically- designed transition state inhibitors are isozyme-specific, with (K(m)/K(i) IU-nucleoside hydrolase)/(K(m)/K(i) IAG-nucleoside hydrolase) values up to 39 000. The mechanism of the differential inhibition is based on the relative leaving group activation and ribosyl-oxocarbenium-forming abilities of these enzymes. In addition to providing isozyme-specific inhibitors, the novel molecules described here have diagnostic value for the nature of the transition states for N-ribohydrolase enzymes.

UR - http://www.scopus.com/inward/record.url?scp=0030956627&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0030956627&partnerID=8YFLogxK

U2 - 10.1021/bi962319v

DO - 10.1021/bi962319v

M3 - Article

VL - 36

SP - 3528

EP - 3534

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 12

ER -