Open AccessProperties of some nuclear nucleases of rat thymocytes and their changes in radiation‐induced apoptosis
Author(s) -
NIKONOVA Larisa V.,
BELETSKY Igor P.,
UMANSKY Samuil R.
Publication year - 1993
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1993.tb18107.x
Subject(s) - nuclease , divalent , micrococcal nuclease , molecular mass , thymocyte , chemistry , size exclusion chromatography , dna , enzyme , gel electrophoresis , sepharose , biochemistry , microbiology and biotechnology , chromatin , biology , organic chemistry , nucleosome , antigen , cd8 , genetics
Three nuclease activites have been found and characterized in rat thymocyte nuclear extracts. A Mn 2+ ‐dependent nuclease is loosely bound to nuclear components and can be extracted with 0.35 M NaCl. The enzyme is activated by Mn 2+ but not by Mg 2+ , Ca 2+ , or both. Its molecular mass is 36–40 kDa when measured by gel filtration and 37 kDa by SDS/PAGE. An acidic nuclease is independent of divalent ions, produces DNA strand breaks with 5′‐OH ends, its molecular mass is about 37 kDa. Two fractions of Ca 2+ /Mn 2+ ‐dependent nuclease, differing in binding to CM‐Sepharose but identical in other respects, are active in the presence of Mn 2+ but can be additionally activated by Ca 2+ . They are inactive in the presence of Mg 2+ or Ca 2+ but cleave DNA in Ca 2+ /Mg 2+ ‐containing medium. The molecular mass of the enzyme is 22 kDa as determined by both gel filtration and electrophoresis. The dependence of nuclease activities on pH, ions, and sulfhydryl reagents is described. Cycloheximide injection to both control and irradiated animals strongly inhibits the activities of Ca 2+ /Mn 2+ ‐dependent nuclease from thymocyte nuclei separated by chromatography on CM‐Sepharose and does not change the activities of Mn 2+ ‐dependent and acidic nucleases. Nuclease activity in thymocyte nuclei from irradiated rats is increased in Ca 2+ /Mg 2+ ‐containing and Ca 2+ /Mn 2+ ‐containing media whereas there is no change in the activity of acidic nuclease. Ca 2+ /Mn 2+ ‐dependent nuclease is extracted from thymocyte nuclei of irradiated rats with 0.35 M NaCl but from control nuclei only with 0.5 M NaCl. Possible reasons of labilization of Ca 2+ /Mn 2+ ‐dependent nuclease binding to the nuclear structures in dying thymocytes are discussed.