Examination of anthrax lethal factor inhibition by siderophores, small hydroxamates, and protamine.
OBJECTIVE
Based on their ability to chelate metals, hydroxamate molecules and siderophores have been successfully used as metalloenzyme inhibitors. As the anthrax toxin lethal factor (LF) is a zinc (Zn)-metallopeptidase, an investigation of the ability of some small non-siderophore hydroxamate compounds, 5 hydroxamate-containing siderophores, and 1 catecholate siderophore was undertaken to determine whether these compounds would inhibit LF. In addition, salmon sperm protamine and ethylenediaminetetraacetic acid were investigated.


METHODS
A spectrophotometric assay of LF activity, based on its reaction with the substrate (Ac-gly-tyr-betaala-arg-arg-arg-arg-arg-arg-arg-arg-val-leu-arg-p-nitroanilide), was used to assess the degree of inhibition of LF by the putative inhibitors. Procedures were implemented to avoid iron contamination of the test solutions and non-ferrated siderophores and hydroxamates were used as potential inhibitors.


RESULTS
The hydroxamate-containing siderophores displayed limited capacities to inhibit LF, as did the low molecular weight hydroxamate compounds. In contrast, the catecholate siderophore enterobactin and the cationic polyamine salmon sperm protamine demonstrated notable inhibition of LF at concentrations ranging from approximately 10 to 200 microM.


CONCLUSIONS
The polyamine salmon sperm protamine which mimics the target site of proteins cleaved by LF, was the most effective inhibitor of the molecules examined, while the small molecule hydroxamates and the hydroxamate siderophores were among the poorest. If chelation of the Zn of LF results in LF inhibition by the molecules examined, it is most likely secondary to binding of the putative inhibitors to the active site of LF.
Thomas
Matthew
M
Department of Biology, Loyola University of Chicago, Chicago, IL Thomas.West@sdstate.edu, United States.
Castignetti
Domenic
D
D000818:Animals; D000881:Anthrax / Q000382:microbiology; D000942:Antigens, Bacterial / Q000737:chemistry / Q000378:metabolism; D001408:Bacillus anthracis / Q000378:metabolism / Q000472:pathogenicity; D001427:Bacterial Toxins / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000378:metabolism; D020134:Catalytic Domain; D006801:Humans; D006877:Hydroxamic Acids / Q000378:metabolism / Q000494:pharmacology*; D008297:Male; D011479:Protamines / Q000378:metabolism / Q000494:pharmacology*; D012474:Salmon; D017262:Siderophores / Q000378:metabolism / Q000494:pharmacology*; D013094:Spermatozoa / Q000737:chemistry
D000942:Antigens, Bacterial; D001427:Bacterial Toxins; D006877:Hydroxamic Acids; D011479:Protamines; D017262:Siderophores; C030325:anthrax toxin