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United States Department of Agriculture

Agricultural Research Service

Title: An avidin-based assay for histone debiotinylase activity in human cell nuclei

Authors
item Chew, Y-C - UNIVERSITY OF NEBRASKA
item Sarath, Gautam
item Zempleni, Janos - UNIVERSITY OF NEBRASKA

Submitted to: Journal of Nutritional Biochemistry
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: September 7, 2006
Publication Date: November 1, 2007
Citation: Chew, Y., Sarath, G., Zempleni, J. 2007. An avidin-based assay for histone debiotinylase activity in human cell nuclei. Journal of Nutritional Biochemistry. 18: 475-481.

Interpretive Summary: In this study we have developed a plate-based assay to study histone debiotinylation. We anticipate that the assay will now allow for the characterization of specific histone debiotinylating enzymes present in the cell nucleus. Discovery of such nuclear enzymes will be of considerable significance in eukaryotic organisms.

Technical Abstract: Covalent binding of biotin to histones participates in heterochromatin formation, cell cycle progression, and the cellular response to DNA breaks. Biotinylation of histones appears to be a reversible process but the identities of enzymes that remove the biotin mark are largely unknown. Our long-term goal is to identify histone debiotinylases in human cells. Here we developed an avidin-based plate assay to quantify histone debiotinylase activities in human cell nuclei. This assay is an essential first step in purifying histone debiotinylases from human cell nuclei. Using this assay we demonstrated that debiotinylation of human histones is a temperature and pH-dependent process, consistent with enzyme catalysis. Studies with purified proteases and competitiors suggested that removal of the biotin mark in histones is mediated by debiotinylases rather than proteases. Activities of histone debiotinylases varied among human tissues: colon = lung > placenta = liver > lymphoid cells. The assay proved useful to monitor activities of semi-purified histone debiotinylases from human body fluids.

Last Modified: 12/22/2014
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