The isolation and partial characterization of low-molecular-weight phosphorylated component of the non-histone proteins of mouse nuclei

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Abstract
After labeling of mouse liver nuclei with [.gamma.-32P]ATP in vitro, 10-20% of the radioactivity incorporated into the saline-soluble nuclear and HAP2 chromatin fractions was located in a low MW component (component 10) with an isoelectric point near 4.5 in urea. By using combinations of ion-exchange chromatography, preparative thin-layer isoelectric focusing and gel filtration, this component was isolated from both nuclear fractions. Recovery from the saline-soluble fraction was poor under conditions that allow endogenous phosphatases to be active. Component 10 was a phosphoprotein on the basis of enzyme-digestion experiments and the detection of phosphoserine and phosphothreonine. The 32P radioactivity did not appear to be associated with phosphorylated basic amino acids. Its MW was determined by gel chromatography and electrophoresis in sodium dodecyl sulphate/polyacrylamide gels as approximately 10,000, and tryptic digestion of the reduced carboxymethylated proteins in urea yielded 2 32P-labeled peptides. No function has yet been assigned to component 10, though its similarity to other low MW acidic proteins is discussed.