Inhibitors of ADP-Ribosylating Bacterial Toxins Based on Oxacarbenium Ion Character at Their Transition States

Guo Chun Zhou, Sapan L. Parikh, Peter C. Tyler, Gary B. Evans, Richard H. Furneaux, Olga V. Zubkova, Paul A. Benjes, Vern L. Schramm

Research output: Contribution to journalArticle

39 Citations (Scopus)

Abstract

The bacterial exotoxins, cholera toxin (CT), pertussis toxin (PT), and diphtheria toxin (DT), interfere with specific host proteins to cause tissue damage for their respective infections. The common toxic mechanism for these agents is mono-ADP-ribosylation of specific amino acids in G, G, and eEF-2 proteins, respectively, by the catalytic A chains of the toxins (CTA, PTA, and DTA). In the absence of acceptor proteins, these toxins also act as NAD+-N-ribosyl hydrolases. The transition-state structures for NAD+ hydrolysis and ADP-ribosylation reactions have oxacarbenium ion character in the ribose. We designed and synthesized analogues of NAD+ to resemble their oxacarbenium ion transition states. Inhibitors with oxacarbenium mimics replacing the NMN-ribosyl group of NAD+ show 200-620-fold increased affinity in the hydrolytic and N-ribosyl transferase reactions catalyzed by CTA. These analogues are also inhibitors for the hydrolysis of NAD+ by PTA with Ki values of 24-40 μM, but bind with similar affinity to the NAD+ substrates. Inhibition of the NAD+ hydrolysis and ADP-ribosyl transferase reactions of DTA gave Ki values from 19 to 48 μM. Catalytic rate enhancements by the bacterial exotoxins are small, and thus transition-state analogues cannot capture large energies of activation. In the cases of DTA and PTA, analogues known to resemble the transition states bind with approximately the same affinity as substrates. Transition-state analogue interrogation of the bacterial toxins indicates that CTA gains catalytic efficiency from modest transition-state stabilization, but DTA and PTA catalyze ADP-ribosyl transferase reactions more from ground-state destabilization. pH dependence of inhibitor action indicated that both neutral and cationic forms of transition-state analogues bind to DTA with similar affinity. The origin of this similarity is proposed to reside in the cationic nature of NAD+ both as substrate and at the transition state.

Original languageEnglish (US)
Pages (from-to)5690-5698
Number of pages9
JournalJournal of the American Chemical Society
Volume126
Issue number18
DOIs
StatePublished - May 12 2004

Fingerprint

Bacterial Toxins
Administrative data processing
NAD
Adenosine Diphosphate
Differential thermal analysis
Ions
Hydrolysis
Proteins
Transferases
Exotoxins
Substrates
Hydrolases
Ground state
Amino acids
Diphtheria Toxin
Stabilization
Chemical activation
Ribose
Poisons
Cholera Toxin

ASJC Scopus subject areas

  • Chemistry(all)

Cite this

Inhibitors of ADP-Ribosylating Bacterial Toxins Based on Oxacarbenium Ion Character at Their Transition States. / Zhou, Guo Chun; Parikh, Sapan L.; Tyler, Peter C.; Evans, Gary B.; Furneaux, Richard H.; Zubkova, Olga V.; Benjes, Paul A.; Schramm, Vern L.

In: Journal of the American Chemical Society, Vol. 126, No. 18, 12.05.2004, p. 5690-5698.

Research output: Contribution to journalArticle

Zhou, Guo Chun ; Parikh, Sapan L. ; Tyler, Peter C. ; Evans, Gary B. ; Furneaux, Richard H. ; Zubkova, Olga V. ; Benjes, Paul A. ; Schramm, Vern L. / Inhibitors of ADP-Ribosylating Bacterial Toxins Based on Oxacarbenium Ion Character at Their Transition States. In: Journal of the American Chemical Society. 2004 ; Vol. 126, No. 18. pp. 5690-5698.
@article{2bc5cfeb5c1e49cb996338c53da1ca33,
title = "Inhibitors of ADP-Ribosylating Bacterial Toxins Based on Oxacarbenium Ion Character at Their Transition States",
abstract = "The bacterial exotoxins, cholera toxin (CT), pertussis toxin (PT), and diphtheria toxin (DT), interfere with specific host proteins to cause tissue damage for their respective infections. The common toxic mechanism for these agents is mono-ADP-ribosylation of specific amino acids in Gsα , Giα, and eEF-2 proteins, respectively, by the catalytic A chains of the toxins (CTA, PTA, and DTA). In the absence of acceptor proteins, these toxins also act as NAD+-N-ribosyl hydrolases. The transition-state structures for NAD+ hydrolysis and ADP-ribosylation reactions have oxacarbenium ion character in the ribose. We designed and synthesized analogues of NAD+ to resemble their oxacarbenium ion transition states. Inhibitors with oxacarbenium mimics replacing the NMN-ribosyl group of NAD+ show 200-620-fold increased affinity in the hydrolytic and N-ribosyl transferase reactions catalyzed by CTA. These analogues are also inhibitors for the hydrolysis of NAD+ by PTA with Ki values of 24-40 μM, but bind with similar affinity to the NAD+ substrates. Inhibition of the NAD+ hydrolysis and ADP-ribosyl transferase reactions of DTA gave Ki values from 19 to 48 μM. Catalytic rate enhancements by the bacterial exotoxins are small, and thus transition-state analogues cannot capture large energies of activation. In the cases of DTA and PTA, analogues known to resemble the transition states bind with approximately the same affinity as substrates. Transition-state analogue interrogation of the bacterial toxins indicates that CTA gains catalytic efficiency from modest transition-state stabilization, but DTA and PTA catalyze ADP-ribosyl transferase reactions more from ground-state destabilization. pH dependence of inhibitor action indicated that both neutral and cationic forms of transition-state analogues bind to DTA with similar affinity. The origin of this similarity is proposed to reside in the cationic nature of NAD+ both as substrate and at the transition state.",
author = "Zhou, {Guo Chun} and Parikh, {Sapan L.} and Tyler, {Peter C.} and Evans, {Gary B.} and Furneaux, {Richard H.} and Zubkova, {Olga V.} and Benjes, {Paul A.} and Schramm, {Vern L.}",
year = "2004",
month = "5",
day = "12",
doi = "10.1021/ja038159+",
language = "English (US)",
volume = "126",
pages = "5690--5698",
journal = "Journal of the American Chemical Society",
issn = "0002-7863",
publisher = "American Chemical Society",
number = "18",

}

TY - JOUR

T1 - Inhibitors of ADP-Ribosylating Bacterial Toxins Based on Oxacarbenium Ion Character at Their Transition States

AU - Zhou, Guo Chun

AU - Parikh, Sapan L.

AU - Tyler, Peter C.

AU - Evans, Gary B.

AU - Furneaux, Richard H.

AU - Zubkova, Olga V.

AU - Benjes, Paul A.

AU - Schramm, Vern L.

PY - 2004/5/12

Y1 - 2004/5/12

N2 - The bacterial exotoxins, cholera toxin (CT), pertussis toxin (PT), and diphtheria toxin (DT), interfere with specific host proteins to cause tissue damage for their respective infections. The common toxic mechanism for these agents is mono-ADP-ribosylation of specific amino acids in Gsα , Giα, and eEF-2 proteins, respectively, by the catalytic A chains of the toxins (CTA, PTA, and DTA). In the absence of acceptor proteins, these toxins also act as NAD+-N-ribosyl hydrolases. The transition-state structures for NAD+ hydrolysis and ADP-ribosylation reactions have oxacarbenium ion character in the ribose. We designed and synthesized analogues of NAD+ to resemble their oxacarbenium ion transition states. Inhibitors with oxacarbenium mimics replacing the NMN-ribosyl group of NAD+ show 200-620-fold increased affinity in the hydrolytic and N-ribosyl transferase reactions catalyzed by CTA. These analogues are also inhibitors for the hydrolysis of NAD+ by PTA with Ki values of 24-40 μM, but bind with similar affinity to the NAD+ substrates. Inhibition of the NAD+ hydrolysis and ADP-ribosyl transferase reactions of DTA gave Ki values from 19 to 48 μM. Catalytic rate enhancements by the bacterial exotoxins are small, and thus transition-state analogues cannot capture large energies of activation. In the cases of DTA and PTA, analogues known to resemble the transition states bind with approximately the same affinity as substrates. Transition-state analogue interrogation of the bacterial toxins indicates that CTA gains catalytic efficiency from modest transition-state stabilization, but DTA and PTA catalyze ADP-ribosyl transferase reactions more from ground-state destabilization. pH dependence of inhibitor action indicated that both neutral and cationic forms of transition-state analogues bind to DTA with similar affinity. The origin of this similarity is proposed to reside in the cationic nature of NAD+ both as substrate and at the transition state.

AB - The bacterial exotoxins, cholera toxin (CT), pertussis toxin (PT), and diphtheria toxin (DT), interfere with specific host proteins to cause tissue damage for their respective infections. The common toxic mechanism for these agents is mono-ADP-ribosylation of specific amino acids in Gsα , Giα, and eEF-2 proteins, respectively, by the catalytic A chains of the toxins (CTA, PTA, and DTA). In the absence of acceptor proteins, these toxins also act as NAD+-N-ribosyl hydrolases. The transition-state structures for NAD+ hydrolysis and ADP-ribosylation reactions have oxacarbenium ion character in the ribose. We designed and synthesized analogues of NAD+ to resemble their oxacarbenium ion transition states. Inhibitors with oxacarbenium mimics replacing the NMN-ribosyl group of NAD+ show 200-620-fold increased affinity in the hydrolytic and N-ribosyl transferase reactions catalyzed by CTA. These analogues are also inhibitors for the hydrolysis of NAD+ by PTA with Ki values of 24-40 μM, but bind with similar affinity to the NAD+ substrates. Inhibition of the NAD+ hydrolysis and ADP-ribosyl transferase reactions of DTA gave Ki values from 19 to 48 μM. Catalytic rate enhancements by the bacterial exotoxins are small, and thus transition-state analogues cannot capture large energies of activation. In the cases of DTA and PTA, analogues known to resemble the transition states bind with approximately the same affinity as substrates. Transition-state analogue interrogation of the bacterial toxins indicates that CTA gains catalytic efficiency from modest transition-state stabilization, but DTA and PTA catalyze ADP-ribosyl transferase reactions more from ground-state destabilization. pH dependence of inhibitor action indicated that both neutral and cationic forms of transition-state analogues bind to DTA with similar affinity. The origin of this similarity is proposed to reside in the cationic nature of NAD+ both as substrate and at the transition state.

UR - http://www.scopus.com/inward/record.url?scp=2442467753&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=2442467753&partnerID=8YFLogxK

U2 - 10.1021/ja038159+

DO - 10.1021/ja038159+

M3 - Article

C2 - 15125661

AN - SCOPUS:2442467753

VL - 126

SP - 5690

EP - 5698

JO - Journal of the American Chemical Society

JF - Journal of the American Chemical Society

SN - 0002-7863

IS - 18

ER -