Analysis of Arg-X proteolytic activity in the supramolecular structures of cell nuclei influenced by inhibitor deacetylation of proteins during the germination of wheat

Abstract

In higher eukaryotes, histone tails and globular domains are subjected to a variety of posttranslational modifications including regulatory proteolysis to create different chromatin structures necessary for the functioning of DNA. The aim of the present work was to detect the regions of activity of Arg-X proteolysis in complexes of histone and non-histone proteins as a potential mechanism affecting the large-scale reorganization of the chromatin. This chromatin reorganization occurs during the germination of wheat germ under conditions of highly acetylated proteins in the cell nuclei. For soaking the seeds and the induction of germination, respectively, distilled water was used as the control, and 0.004 mM sodium butyrate as the test. Cell nuclei were isolated from germs, cleared, and then the nucleoplasm, chromatin and the nuclear matrix were extracted by increasing the ionic strength of the solution. From isolated supra-molecular structures, non-histone proteins were separated from histones using ion exchange chromatography. The Arg-X proteolytic activity was assessed by cleavage of Arg-X bonds in the arginine-enriched protein protamine in all nuclear fractions. It is possible that decreasing in histone turnover due to the antiproliferative effect of the deacetylase inhibitor (sodium butyrate) is due to decreasing of proteolytic activity in histone and non-histone proteins associated with chromatin, suggesting the possible role of histone proteinases in histone metabolism. These results also enabled us to detect the regions of activity of Arg-X proteolysis in the non-histone and histone blocks. It also provided information on the specific tissues in which proteolysis activity is observed during wheat germ germination under normal conditions, as well as under conditions of highly acetylated proteins in the cell nuclei. These results provide an example of the proteinase network formed in the supra-molecular structures of cell nuclei.