Submitted to: Journal of Nutritional Biochemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/8/2007
Publication Date: 12/1/2008
Citation: Kobza, K., Chaiseeda, K., Sarath, G., Takacs, J.M., Zempleni, J. 2008. Biotinyl-methyl 4-(amidomethyl) benzoate is a competitive inhibitor of human biotinidase. Journal of Nutritional Biochemistry. Vol 19: 826-832. Interpretive Summary: Histones are DNA-binding proteins. Modification of histones by the vitamin, biotin can cause changes in gene expression. The enzyme biotinidase can participate in the addition or removal of biotin to histones. In this study, new chemicals were evaluated for the ability to inhibit biotinidase in-vitro. One compound, biotinyl-methyl 4-(amidomethyl) benzoate was found to cause significant inhibition of biotinidase activity. Finding good inhibitors for biotinidase could have future clinical impact in the treatment of disease states linked to the level of histone biotinylation.
Technical Abstract: Posttranslational modification of histones by biotinylation can be catalyzed by both biotinidase (BTD) and holocarboxylase synthetase (HCS). Biotinylation of histones is an important epigenetic mechanism to regulate gene expression, DNA repair, and chromatin remodeling. The role of BTD in histone biotinylation is somewhat ambiguous, given that BTD also catalyzes removal of the biotin tag from histones. Here, we sought to develop BTD inhibitors for future studies of the role of BTD in altering chromatin structure. We adopted an existing colorimetric BTD assay for use in a novel 96-well plate format to permit high-throughput screening of potential inhibitors. Biotin analogs were chemically synthesized and tested for their ability to inhibit human BTD. Seven of these compounds inhibited BTD by 26% to 80%. Biotinyl-methyl 4-(amidomethyl) benzoate had the largest effect on BTD, causing an 80% inhibition at 1 mM concentration. Enzyme kinetics studies were conducted to determine Vmax, Km, and Ki for the seven inhibitors; kinetics were consistent with the hypothesis that biotinyl-methyl 4-(amidomethyl) benzoate and the other compounds acted by competitive inhibition of BTD. Finally, biotinyl-methyl 4-(amidomethyl) benzoate did not affect biotin transport in human cells, suggesting specificity in regard to biotin-related processes.