Formation in vitro of the pTP-dCMP initiation complex of human adenovirus type 12

• Published in: Journal of general virology 1991-02, Vol.72 (2), p.427-430
• Main Authors: Chowrira, Bharat M, Zhao, Jiansheng, Lucher, Lynne A
• Format: Article
• Language: English
• Subjects: Adenoviruses, Human - metabolism; Amino Acid Sequence; Cytidine Triphosphate - metabolism; Deoxycytidine Monophosphate - metabolism; Humans; Immunoblotting; KB Cells; Molecular Sequence Data; Viral Proteins - metabolism; Virus Replication
• ISSN: 0022-1317; 1465-2099
• DOI: 10.1099/0022-1317-72-2-427

Microbiology Group, Department of Biological Sciences, Illinois State University, Normal, Illinois 61761, U.S.A. We report the covalent addition of [ 32 P]dCMP to a protein from group A adenovirus 12 (Ad12)-infected human (KB) cells in vitro , using crude extracts. Synthesis of the 60K protein-dCMP complex required a DNA template containing a terminally located adenovirus replication origin; the protein-dCMP bond was alkali-labile but acid-stable. We therefore conclude that this product is the Ad12 terminal protein precursor (pTP)-dCMP initiation complex for DNA replication. Synthesis of Ad12 pTP-dCMP was specific for dCTP but was stimulated by dATP. In contrast to Ad2, the Ad12 initiation reaction required ATP. Antipeptide antiserum targeted to Ad DNA polymerase inhibited Ad12 pTP-dCMP synthesis in vitro , providing evidence that Ad DNA polymerase catalyses dCMP addition to pTP during initiation. Present address: Department of Microbiology and Molecular Genetics, University of Vermont College of Medicine, Burlington, Vermont 05405, U.S.A. Received 27 June 1990; accepted 16 October 1990.