Phosphoproteins of the Cell Nucleus: Metabolism and Characterization

Author

Lewis Joel Kleinsmith

Date of Award

1968

Document Type

Thesis

Degree Name

Doctor of Philosophy (PhD)

RU Laboratory

Mirsky Laboratory

Abstract

This thesis is concerned with the chemistry and metabolism of the phosphorylated proteins which are found localized in cell nuclei. The studies focus first on phosphoprotein metabolism as it occurs in the intact nucleus under varying conditions, and then on the chemistry and enzymology of a phosphoprotein fraction which has been extracted and purified from isolated nuclei. The metabolism of phosphoproteins was studied in isolated calf thymus nuclei employing P³² -orthophosphate as a tracer. These isolated nuclei incorporate this isotope into nuclear proteins, which yield phosphoserine and phosphothreonine on hydrolysis. Protein phosphorylation is energy dependent, and occurs after the polypeptide chain has been completed. The phosphate groups, once incorporated into the protein, are not stable, but are subject to a very rapid "turnover" reaction. This "turnover" reaction in the nuclei also appears to be energy dependent. The bulk of phosphate incorporation in these nuclei is into non-histone protein, but small amounts of radioactive serine phosphate can be detected in highly purified histone fractions. Since nuclear phosphoproteins have been suggested to play a role in regulating RNA synthesis, experiments were performed to determine whether the rate of protein phosphorylation in nuclei increases when gene activity is stimulated. Human lymphocytes treated with phytohemagglutinin undergo extensive gene activation, as evidenced by augmented synthesis of RNA. This activation is preceded by an early stimulation in the rate of phosphorylation and dephosphorylation of nuclear proteins. This finding is consistent with the hypothesized role of phosphoproteins in the modification of chromatin structure and in modulation of the template activity of DNA in vivo.

Comments

A thesis submitted to the Faculty of The Rockefeller University in partial fulfillment of the requirements for the degree of Doctor of Philosophy

Recommended Citation

Kleinsmith, Lewis Joel, "Phosphoproteins of the Cell Nucleus: Metabolism and Characterization" (1968). Student Theses and Dissertations. 582.
https://digitalcommons.rockefeller.edu/student_theses_and_dissertations/582