H3-specific nucleohistone kinase of bovine thymus chromatin. Purification, characterization, and specificity for threonine residue 3.
Bovine thymus chromatin contains a cAMP-independent histone kinase which is entirely specific for a single site on H3 whether the histone substrate is soluble or associated with DNA in chromatin (Shoemaker, C. B., and Chalkley, R. (1978) J. Biol. Chem. 253, 5802--5807). The H3-kinase has been extracted, purified 2000-fold and extensively characterized. The purified enzyme produces a single band upon neutral gel electrophoresis and two distinct bands of 21,000 and 23,000 daltons upon sodium dodecyl sulfate-gel electrophoresis. Following subcellular fractionation, most or all of the enzymatic activity is associated with chromatin. Soluble histone inactivates H3-kinase after short incubations while a chromatin substrate permits the enzyme to remain active until H3 is fully phosphorylated. Assay conditions have been optimized in terms of pH and several cofactor concentrations. Optimal MgCl2 concentration occurs at 50 mM, while for MnCl2 the optimum is 300 microM. H3-kinase has a molecular weight of 38,000 as estimated by exclusion chromatography. The Km for ATP is 160 +/- 23 microM. The enzyme displays extraordinary substrate specificity for H3 histone as no other thymus protein has been observed as a substrate. Phosphorylation of H3 occurs at threonine residue 3.
D000818:Animals; D002417:Cattle; D002843:Chromatin / Q000201:enzymology*; D007700:Kinetics; D008970:Molecular Weight; D006656:Protamine Kinase / Q000302:isolation & purification / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism*; D013379:Substrate Specificity; D013912:Threonine; D013950:Thymus Gland / Q000201:enzymology*
D002843:Chromatin; D013912:Threonine; D011494:Protein Kinases; D006656:Protamine Kinase