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Laser cross-linking of proteins to nucleic acids. II. Interactions of the bacteriophage T4 DNA replication polymerase accessory proteins complex with DNA. 1993

J W Hockensmith, and W L Kubasek, and E M Evertsz, and L D Mesner, and P H von Hippel
Institute of Molecular Biology, University of Oregon, Eugene 97403.

In this paper we examine the interactions of the polymerase accessory proteins subassembly of the bacteriophage T4 DNA replication complex, using single-pulse ultraviolet laser excitation to induce protein-nucleic acid cross-links. The laser-induced cross-linking permits effective "freezing" of the instantaneous equilibrium state of the complex and thus provides a mechanism to dissect the individual protein-nucleic acid interactions involved in complex assembly. We find that the binding of the gene 44, 62, and 45 proteins is dependent not only on the presence of each of the other proteins, but also on the presence of adenine nucleotide cofactors. We find that the nonhydrolyzable analogs of ATP often behave more like ADP than ATP in these experiments. Gene 45 protein is able to induce an increase in cross-linking of the gp44/62 complex to nucleic acids, and this increased cross-linking correlates with changes in the apparent Km of the gp44/62 complex for polynucleotides and with changes in Vmax during ATP hydrolysis. Our results suggest that the enhanced DNA binding is predominately through the gene 62 protein and not the ATPase catalytic subunit (gene 44 protein). Thus the gene 62 protein seems to play an integral role in gp45-mediated enhancement of the ATP hydrolytic activity of gp44. These results are summarized and integrated in the form of a model for the multiple interactions of the accessory proteins with DNA and one another in the presence of mononucleotide cofactors and substrates.

UI MeSH Term Description Entries
D007834 Lasers An optical source that emits photons in a coherent beam. Light Amplification by Stimulated Emission of Radiation (LASER) is brought about using devices that transform light of varying frequencies into a single intense, nearly nondivergent beam of monochromatic radiation. Lasers operate in the infrared, visible, ultraviolet, or X-ray regions of the spectrum. Masers,Continuous Wave Lasers,Pulsed Lasers,Q-Switched Lasers,Continuous Wave Laser,Laser,Laser, Continuous Wave,Laser, Pulsed,Laser, Q-Switched,Lasers, Continuous Wave,Lasers, Pulsed,Lasers, Q-Switched,Maser,Pulsed Laser,Q Switched Lasers,Q-Switched Laser
D004259 DNA-Directed DNA Polymerase DNA-dependent DNA polymerases found in bacteria, animal and plant cells. During the replication process, these enzymes catalyze the addition of deoxyribonucleotide residues to the end of a DNA strand in the presence of DNA as template-primer. They also possess exonuclease activity and therefore function in DNA repair. DNA Polymerase,DNA Polymerases,DNA-Dependent DNA Polymerases,DNA Polymerase N3,DNA Dependent DNA Polymerases,DNA Directed DNA Polymerase,DNA Polymerase, DNA-Directed,DNA Polymerases, DNA-Dependent,Polymerase N3, DNA,Polymerase, DNA,Polymerase, DNA-Directed DNA,Polymerases, DNA,Polymerases, DNA-Dependent DNA
D004261 DNA Replication The process by which a DNA molecule is duplicated. Autonomous Replication,Replication, Autonomous,Autonomous Replications,DNA Replications,Replication, DNA,Replications, Autonomous,Replications, DNA
D004279 DNA, Viral Deoxyribonucleic acid that makes up the genetic material of viruses. Viral DNA
D000244 Adenosine Diphosphate Adenosine 5'-(trihydrogen diphosphate). An adenine nucleotide containing two phosphate groups esterified to the sugar moiety at the 5'-position. ADP,Adenosine Pyrophosphate,Magnesium ADP,MgADP,Adenosine 5'-Pyrophosphate,5'-Pyrophosphate, Adenosine,ADP, Magnesium,Adenosine 5' Pyrophosphate,Diphosphate, Adenosine,Pyrophosphate, Adenosine
D000255 Adenosine Triphosphate An adenine nucleotide containing three phosphate groups esterified to the sugar moiety. In addition to its crucial roles in metabolism adenosine triphosphate is a neurotransmitter. ATP,Adenosine Triphosphate, Calcium Salt,Adenosine Triphosphate, Chromium Salt,Adenosine Triphosphate, Magnesium Salt,Adenosine Triphosphate, Manganese Salt,Adenylpyrophosphate,CaATP,CrATP,Manganese Adenosine Triphosphate,MgATP,MnATP,ATP-MgCl2,Adenosine Triphosphate, Chromium Ammonium Salt,Adenosine Triphosphate, Magnesium Chloride,Atriphos,Chromium Adenosine Triphosphate,Cr(H2O)4 ATP,Magnesium Adenosine Triphosphate,Striadyne,ATP MgCl2
D013604 T-Phages A series of 7 virulent phages which infect E. coli. The T-even phages T2, T4; (BACTERIOPHAGE T4), and T6, and the phage T5 are called "autonomously virulent" because they cause cessation of all bacterial metabolism on infection. Phages T1, T3; (BACTERIOPHAGE T3), and T7; (BACTERIOPHAGE T7) are called "dependent virulent" because they depend on continued bacterial metabolism during the lytic cycle. The T-even phages contain 5-hydroxymethylcytosine in place of ordinary cytosine in their DNA. Bacteriophages T,Coliphages T,Phages T,T Phages,T-Phage
D013698 Templates, Genetic Macromolecular molds for the synthesis of complementary macromolecules, as in DNA REPLICATION; GENETIC TRANSCRIPTION of DNA to RNA, and GENETIC TRANSLATION of RNA into POLYPEPTIDES. Genetic Template,Genetic Templates,Template, Genetic
D014764 Viral Proteins Proteins found in any species of virus. Gene Products, Viral,Viral Gene Products,Viral Gene Proteins,Viral Protein,Protein, Viral,Proteins, Viral

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