Structure–function analysis of water-soluble inhibitors of the catalytic domain of exotoxin A from Pseudomonas aeruginosa
Open Access
- 24 January 2005
- journal article
- Published by Portland Press Ltd. in Biochemical Journal
- Vol. 385 (3), 667-675
- https://doi.org/10.1042/bj20041480
Abstract
The mono-ADPRT (mono-ADP-ribosyltransferase), Pseudomonas aeruginosa ETA (exotoxin A), catalyses the transfer of ADP-ribose from NAD+ to its protein substrate. A series of water-soluble compounds that structurally mimic the nicotinamide moiety of NAD+ was investigated for their inhibition of the catalytic domain of ETA. The importance of an amide locked into a hetero-ring structure and a core hetero-ring system that is planar was a trend evident by the IC50 values. Also, the weaker inhibitors have core ring structures that are less planar and thus more flexible. One of the most potent inhibitors, PJ34, was further characterized and shown to exhibit competitive inhibition with an inhibition constant Ki of 140 nM. We also report the crystal structure of the catalytic domain of ETA in complex with PJ34, the first example of a mono-ADPRT in complex with an inhibitor. The 2.1 Å (1 Å=0.1 nm) resolution structure revealed that PJ34 is bound within the nicotinamide-binding pocket and forms stabilizing hydrogen bonds with the main chain of Gly-441 and to the side-chain oxygen of Gln-485, a member of a proposed catalytic loop. Structural comparison of this inhibitor complex with diphtheria toxin (a mono-ADPRT) and with PARPs [poly(ADP-ribose) polymerases] shows similarity of the catalytic residues; however, a loop similar to that found in ETA is present in diphtheria toxin but not in PARP. The present study provides insight into the important features required for inhibitors that mimic NAD+ and their binding to the mono-ADPRT family of toxins.Keywords
This publication has 42 references indexed in Scilit:
- Transition State Structure for ADP-Ribosylation of Eukaryotic Elongation Factor 2 Catalyzed by Diphtheria ToxinBiochemistry, 2004
- Poly(ADP-Ribose) Polymerase Inhibition Reduces Reperfusion Injury After Heart TransplantationCirculation Research, 2002
- Characterization of Competitive Inhibitors for the Transferase Activity of Pseudomonas aeruginosa Exotoxin AJournal of Enzyme Inhibition and Medicinal Chemistry, 2002
- A Catalytic Loop within Pseudomonas aeruginosa Exotoxin A Modulates Its Transferase ActivityJournal of Biological Chemistry, 2001
- GPI 6150 Prevents H2O2 Cytotoxicity by Inhibiting Poly(ADP-ribose) PolymeraseBiochemical and Biophysical Research Communications, 2000
- http://www.cdc.gov/ncidod/eid/vol4no4/vandelden.htmEmerging Infectious Diseases, 1998
- Unusual conformation of nicotinamide adenine dinucleotide (NAD) bound to diphtheria toxin: A comparison with NAD bound to the oxidoreductase enzymesProtein Science, 1997
- SWISS‐MODEL and the Swiss‐Pdb Viewer: An environment for comparative protein modelingElectrophoresis, 1997
- Mutations in the Elongation Factor 2 Gene Which Confer Resistance to Diphtheria Toxin and Pseudomonas Exotoxin APublished by Elsevier BV ,1995
- Pseudomonas aeruginosa exotoxin A: effects of mutating tyrosine-470 and tyrosine-481 to phenylalanineBiochemistry, 1988