Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Botulinum neurotoxin A protease: discovery of natural product exosite inhibitors.

ABSTRACT: A new mechanistic class of BoNT/A zinc metalloprotease inhibitors, from Echinacea, exemplified by the natural product d-chicoric acid (I1) is disclosed. A detailed evaluation of chicoric acid's mechanism of inhibition reveals that the inhibitor binds to an exosite, displays noncompetitive partial inhibition, and is synergistic with a competitive active site inhibitor when used in combination. Other components found in Echinacea, I3 and I4, were also inhibitors of the protease.

SUBMITTER: Silhar P

PROVIDER: S-EPMC2832098 | biostudies-literature | 2010 Mar

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Publications

Botulinum neurotoxin A protease: discovery of natural product exosite inhibitors.

Silhár Peter P Capková Katerina K Salzameda Nicholas T NT Barbieri Joseph T JT Hixon Mark S MS Janda Kim D KD

Journal of the American Chemical Society 20100301 9

A new mechanistic class of BoNT/A zinc metalloprotease inhibitors, from Echinacea, exemplified by the natural product d-chicoric acid (I1) is disclosed. A detailed evaluation of chicoric acid's mechanism of inhibition reveals that the inhibitor binds to an exosite, displays noncompetitive partial inhibition, and is synergistic with a competitive active site inhibitor when used in combination. Other components found in Echinacea, I3 and I4, were also inhibitors of the protease. ...[more]

PMID: 20158239

Similar Datasets

Structural insight into exosite binding and discovery of novel exosite inhibitors of botulinum neurotoxin serotype A through in silico screening.

Project description:Botulinum neurotoxin serotype A (BoNT/A) is the most lethal toxin among the Tier 1 Select Agents. Development of potent and selective small molecule inhibitors against BoNT/A zinc metalloprotease remains a challenging problem due to its exceptionally large substrate binding surface and conformational plasticity. The exosites of the catalytic domain of BoNT/A are intriguing alternative sites for small molecule intervention, but their suitability for inhibitor design remains largely unexplored. In this study, we employed two recently identified exosite inhibitors, D-chicoric acid and lomofungin, to probe the structural features of the exosites and molecular mechanisms of synergistic inhibition. The results showed that D-chicoric acid favors binding at the ?-exosite, whereas lomofungin preferentially binds at the ?-exosite by mimicking the substrate ?-sheet binding interaction. Molecular dynamics simulations and binding interaction analysis of the exosite inhibitors with BoNT/A revealed key elements and hotspots that likely contribute to the inhibitor binding and synergistic inhibition. Finally, we performed database virtual screening for novel inhibitors of BoNT/A targeting the exosites. Hits C1 and C2 showed non-competitive inhibition and likely target the ?- and ?-exosites, respectively. The identified exosite inhibitors may provide novel candidates for structure-based development of therapeutics against BoNT/A intoxication.

| S-EPMC4138512 | biostudies-literature

Identification of a Natural Product Antagonist against the Botulinum Neurotoxin Light Chain Protease.

Project description:Botulinum neurotoxins (BoNTs) are the etiological agents responsible for botulism, a disease characterized by peripheral neuromuscular blockade and a characteristic flaccid paralysis of humans. BoNTs are the most lethal known poisons affecting humans and has been recognized as a potential bioterrorist threat. Current treatments for botulinum poisoning are predominately prophylactic in nature relying on passive immunization with antitoxins. Inhibition of the BoNT light chain metalloprotease (LC) has emerged as a new therapeutic strategy for the treatment of botulism that may provide an effective post-exposure remedy. A high-throughput screening effort against the light chain of BoNT serotype A (LC/A) was conducted with the John Hopkins Clinical Compound Library comprised of over 1,500 existing drugs. Lomofungin, a natural product first isolated in the late 1960's, was identified as an inhibitor of LC/A, displaying classical noncompetitive inhibition kinetics with a K(i) of 6.7 ± 0.7 µM. Inhibitor combination studies reveal that lomofungin binding is nonmutually exclusive (synergistic). The inhibition profile of lomofungin has been delineated by the use of both an active site inhibitor, 2,4-dichlorocinnamic hydroxamate, and a noncompetitive inhibitor d-chicoric acid; the mechanistic implications of these observations are discussed. Lastly, cellular efficacy was investigated using a rat primary cell model which demonstrated that lomofungin can protect against SNAP-25 cleavage, the intracellular protein target of LC/A.

| S-EPMC2955888 | biostudies-literature

Examination of ?-exosite inhibitors against Botulinum neurotoxin A protease through structure-activity relationship studies of chicoric acid.

Project description:Botulinum neurotoxins (BoNT) are among the most toxic known substances and currently there are no effective treatments for intraneuronal BoNT intoxication. Chicoric acid (ChA) was previously reported as a BoNT/A inhibitor that binds to the enzyme's ?-exosite. Herein, we report the synthesis and structure-activity relationships (SARs) of a series of ChA derivatives, which revealed essential binding interactions between ChA and BoNT/A. Moreover, several ChA-based inhibitors with improved potency against the BoNT/A were discovered.

| S-EPMC5667901 | biostudies-other

Picolinic acids as ?-exosite inhibitors of botulinum neurotoxin A light chain.

Project description:In developing small-molecule inhibitors of botulinum neurotoxin serotype A light chain (BoNT/A LC), substituted picolinic acids were identified. Extensive investigation into the SAR of the picolinic acid scaffold revealed 5-(1-butyl-4-chloro-1H-indol-2-yl)picolinic acid (CBIP), which possessed low micromolar activity against BoNT/A. Kinetic and docking studies demonstrated binding of CBIP to the ?-exosite: a largely unexplored site on the LC that holds therapeutic relevance for botulism treatment.

| S-EPMC5085802 | biostudies-literature

Reexamining hydroxamate inhibitors of botulinum neurotoxin serotype A: extending towards the ?-exosite.

Project description:Botulinum neurotoxins (BoNTs) are the most toxic proteins known to man, exposure to which results in flaccid paralysis. Given their extreme potency, these proteins have become studied as possible weapons of bioterrorism; however, effective treatments that function after intoxication have not progressed to the clinic. Here, we have reexamined one of the most effective inhibitors, 2,4-dichlorocinnamyl hydroxamate, in the context of the known plasticity of the BoNT/A light chain metalloprotease. Our studies have shown that modifications of this compound are tolerated and result in improved inhibitors, with the best compound having an IC(50) of 0.23 ?M. Given the inconsistency of structure-activity relationship trends observed across similar compounds, this data argues for caution in extrapolating across structural series.

| S-EPMC3352981 | biostudies-literature

A cross-over inhibitor of the botulinum neurotoxin light chain B: a natural product implicating an exosite mechanism of action.

Project description:Clostridium botulinum produces the most lethal toxins known to man, as such they are high risk terrorist threats, and alarmingly there is no approved therapeutic. We report the first cross-over small molecule inhibitor of these neurotoxins and propose a mechanism by which it may impart its inhibitory activity.

| S-EPMC3065946 | biostudies-literature

Synthesis and biological evaluation of botulinum neurotoxin a protease inhibitors.

Project description:NSC 240898 was previously identified as a botulinum neurotoxin A light chain (BoNT/A LC) endopeptidase inhibitor by screening the National Cancer Institute Open Repository diversity set. Two types of analogues have been synthesized and shown to inhibit BoNT/A LC in a FRET-based enzyme assay, with confirmation in an HPLC-based assay. These two series of compounds have also been evaluated for inhibition of anthrax lethal factor (LF), an unrelated metalloprotease, to examine enzyme specificity of the BoNT/A LC inhibition. The most potent inhibitor against BoNT/A LC in these two series is compound 12 (IC(50) = 2.5 microM, FRET assay), which is 4.4-fold more potent than the lead structure and 11.2-fold more selective for BoNT/A LC versus the anthrax LF metalloproteinase. Structure-activity relationship studies have revealed structural features important to potency and enzyme specificity.

| S-EPMC2841792 | biostudies-literature

Crystal structure of Clostridium botulinum neurotoxin protease in a product-bound state: Evidence for noncanonical zinc protease activity.

Project description:Clostridium botulinum neurotoxins (BoNTs), the most potent toxins known, disrupt neurotransmission through proteolysis of proteins involved in neuroexocytosis. The light chains of BoNTs are unique zinc proteases that have stringent substrate specificity and require exceptionally long substrates. We have determined the crystal structure of the protease domain from BoNT serotype A (BoNT/A). The structure reveals a homodimer in a product-bound state, with loop F242-V257 from each monomer deeply buried in its partner's catalytic site. The loop, which acts as a substrate, is oriented in reverse of the canonical direction for other zinc proteases. The Y249-Y250 peptide bond of the substrate loop is hydrolyzed, leaving the Y249 product carboxylate coordinated to the catalytic zinc. From the crystal structure of the BoNT/A protease, detailed models of noncanonical binding and proteolysis can be derived which we propose are also consistent with BoNT/A binding and proteolysis of natural substrate synaptosome-associated protein of 25 kDa (SNAP-25). The proposed BoNT/A substrate-binding mode and catalytic mechanism are markedly different from those previously proposed for the BoNT serotype B.

| S-EPMC406437 | biostudies-literature

Tyrosine phosphorylation of botulinum neurotoxin protease domains.

Project description:Botulinum neurotoxins are most potent of all toxins. Their N-terminal light chain domain (Lc) translocates into peripheral cholinergic neurons to exert its endoproteolytic action leading to muscle paralysis. Therapeutic development against these toxins is a major challenge due to their in vitro and in vivo structural differences. Although three-dimensional structures and reaction mechanisms are very similar, the seven serotypes designated A through G vastly vary in their intracellular catalytic stability. To investigate if protein phosphorylation could account for this difference, we employed Src-catalyzed tyrosine phosphorylation of the Lc of six serotypes namely LcA, LcB, LcC1, LcD, LcE, and LcG. Very little phosphorylation was observed with LcD and LcE but LcA, LcB, and LcG were maximally phosphorylated by Src. Phosphorylation of LcA, LcB, and LcG did not affect their secondary and tertiary structures and thermostability significantly. Phosphorylation of Y250 and Y251 made LcA resistant to autocatalysis and drastically reduced its k(cat)/K(m) for catalysis. A tyrosine residue present near the essential cysteine at the C-terminal tail of LcA, LcB, and LcG was readily phosphorylated in vitro. Inclusion of a competitive inhibitor protected Y426 of LcA from phosphorylation, shedding light on the role of the C-terminus in the enzyme's substrate or product binding.

| S-EPMC3366388 | biostudies-literature

Synthesis and structure-activity relationships of second-generation hydroxamate botulinum neurotoxin A protease inhibitors.

Project description:Botulinum neurotoxins are the most toxic proteins currently known. Based on a recently identified potent lead structure, 2,4-dichlorocinnamic acid hydroxamate, herein we report on the structure-activity relationship of a series of hydroxamate BoNT/A inhibitors. Among them, 2-bromo-4-chlorocinnamic acid hydroxamate, 2-methyl-4-chlorocinnamic acid hydroxamate, and 2-trifluoromethyl-4-chlorocinnamic acid hydroxamate displayed comparable inhibitory activity to that of the lead structure.

| S-EPMC2170532 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data