Enzymatic activities associated with a purified simian virus 40 T antigen-related protein

Tjian, R., Robbins, A. (February 1979) Enzymatic activities associated with a purified simian virus 40 T antigen-related protein. Proc Natl Acad Sci U S A, 76 (2). pp. 610-4. ISSN 0027-8424 (Print)0027-8424 (Linking)

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URL: http://www.ncbi.nlm.nih.gov/pubmed/218212

Abstract

A protein antigenically related to the simian virus (SV 40) A gene product has been purified to near homogeneity from cells infected with the adenovirus-SV 40 hybrid virus Ad2(+)D2 and shown to contain ATPase (ATP phosphohydrolase, EC 3.6.1.3) and protein kinase (ATP:phosphotransferase, EC 2.7.1.37) activity. Both enzymatic activities copurify with the protein through six stages including one gel filtration column, two ion exchange columns, and a heparin affinity column. Analogous fractions from extracts of cells uninfected or infected with adenovirus 2 alone do not contain these enzymatic activities. The D2 hybrid protein resolves into two forms (I and II) during ion exchange chromatography. Form I, the major species (85%) of the D2 hybrid protein, elutes from DEAE-Sephadex in 0.37 M NaCl and is able to catalyze the hydrolysis of ATP to ADP + P(i) at a rate of 3 mumol/hr per mg. The remaining 10-15% of the D2 hybrid protein consists of form II which elutes from DEAE-Sephadex in 0.29 M NaCl and is able to hydrolyze ATP as well as to incorporate phosphorus from ATP into either the D2 hybrid protein itself or other protein acceptors such as phosvitin. Although both forms are able to bind DNA, the ATPase activity of form I cosediments with SV 40 DNA more efficiently than does the protein kinase activity of form II during glycerol gradient centrifugation. The ATPase activity of form I is efficiently inhibited by addition of anti-T gamma globulin to the reaction mixture whereas control gamma globulin has no effect. Similarly, the phosphorylation of the D2 hybrid protein by form II is inhibited by anti-T gamma globulin. By contrast, phosphorylation of phosvitin is specifically inhibited by antibody only when the immune complex is removed from the reaction mixture. Thus, it appears likely that one and possibly two enzymatic activities are carried out by the D2 hybrid protein. These findings are discussed in terms of mechanisms of SV 40 DNA replication and virally induced transformation.

Item Type: Paper
Uncontrolled Keywords: Adenosine Triphosphatases/metabolism Adenoviruses, Human/metabolism Antigen-Antibody Reactions Antigens, Neoplasm/*metabolism DNA Replication Molecular Weight Protein Kinases/metabolism Simian virus 40/*enzymology/immunology Viral Proteins/isolation & purification/metabolism Virus Replication
Subjects: bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > DNA replication
organism description > virus > adenovirus
organism description > virus > SV40
CSHL Authors:
Communities: CSHL labs
Depositing User: Matt Covey
Date: February 1979
Date Deposited: 06 May 2016 20:57
Last Modified: 09 Nov 2017 21:01
PMCID: PMC382998
Related URLs:
URI: http://repository.cshl.edu/id/eprint/32714

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