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Kinetic parameters of the translocation of bacteriophage T4 gene 41 protein helicase on single-stranded DNA. 1994

M C Young, and D E Schultz, and D Ring, and P H von Hippel
Institute of Molecular Biology, University of Oregon, Eugene 97403.

The bacteriophage T4 gene 41 helicase protein (gp41) carries a single-stranded DNA-dependent ATPase activity that is essential to its helicase activity. This ATPase activity can be stimulated by a wide variety of single-stranded DNA cofactors, including homo-oligomers and homopolymers 8 to approximately 10,000 nucleotide residues in length, and by natural single-stranded DNA, such as bacteriophage M13 DNA. The steady-state ATPase activity of gp41 on single-stranded homopolymeric cofactors is dependent on the length of the cofactor, in that the kinetic parameters Vmax and K(act) (or KmDNA) have a characteristic length dependence. Vmax values for different DNA lengths show a hyperbolic dependence on DNA length, while K(act) values are independent of DNA lengths exceeding approximately 20 nucleotide residues. Use of the detailed theoretical analysis developed in the preceding paper reveals that: (1) these results support the earlier proposal that gp41 translocates on single-stranded DNA in an ATP-dependent manner; (2) translocation is undirectional; (3) translocation is processive to an extent that depends on the base composition of the DNA employed, with the average distance translocated per binding event ranging from 60 to 700 nucleotide residues; and (4) the detailed translocation mechanism of gp41 includes an obligatory slow step before or after the ATP-driven translocation process. Defined lengths of natural and homopolymer single-stranded DNA have also been created as gaps of known length distribution between clusters of gene 32 protein (gp32) bound along long single-stranded DNA molecules. ATPase data obtained with cofactors of this type also show unidirectional ATP-driven translocation of gp41 on both natural and homopolymeric single-stranded DNA. Direct binding studies of gp41 to short dT oligomers reveal two further features of the interaction of gp41 to single-stranded DNA: (1) nucleoside triphosphate binding is necessary for the formation of stable gp41-ssDNA complexes; and (2) the DNA binding site size of gp41 is between 12 and 20 nucleotide residues per protein monomer. Possible translocation mechanisms for gp41 are discussed within the context of these results.

UI MeSH Term Description Entries
D007700 Kinetics The rate dynamics in chemical or physical systems.
D008433 Mathematics The deductive study of shape, quantity, and dependence. (From McGraw-Hill Dictionary of Scientific and Technical Terms, 6th ed) Mathematic
D008962 Models, Theoretical Theoretical representations that simulate the behavior or activity of systems, processes, or phenomena. They include the use of mathematical equations, computers, and other electronic equipment. Experimental Model,Experimental Models,Mathematical Model,Model, Experimental,Models (Theoretical),Models, Experimental,Models, Theoretic,Theoretical Study,Mathematical Models,Model (Theoretical),Model, Mathematical,Model, Theoretical,Models, Mathematical,Studies, Theoretical,Study, Theoretical,Theoretical Model,Theoretical Models,Theoretical Studies
D004265 DNA Helicases Proteins that catalyze the unwinding of duplex DNA during replication by binding cooperatively to single-stranded regions of DNA or to short regions of duplex DNA that are undergoing transient opening. In addition, DNA helicases are DNA-dependent ATPases that harness the free energy of ATP hydrolysis to translocate DNA strands. ATP-Dependent DNA Helicase,DNA Helicase,DNA Unwinding Protein,DNA Unwinding Proteins,ATP-Dependent DNA Helicases,DNA Helicase A,DNA Helicase E,DNA Helicase II,DNA Helicase III,ATP Dependent DNA Helicase,ATP Dependent DNA Helicases,DNA Helicase, ATP-Dependent,DNA Helicases, ATP-Dependent,Helicase, ATP-Dependent DNA,Helicase, DNA,Helicases, ATP-Dependent DNA,Helicases, DNA,Protein, DNA Unwinding,Unwinding Protein, DNA,Unwinding Proteins, DNA
D004277 DNA, Single-Stranded A single chain of deoxyribonucleotides that occurs in some bacteria and viruses. It usually exists as a covalently closed circle. Single-Stranded DNA,DNA, Single Stranded,Single Stranded DNA
D004926 Escherichia coli A species of gram-negative, facultatively anaerobic, rod-shaped bacteria (GRAM-NEGATIVE FACULTATIVELY ANAEROBIC RODS) commonly found in the lower part of the intestine of warm-blooded animals. It is usually nonpathogenic, but some strains are known to produce DIARRHEA and pyogenic infections. Pathogenic strains (virotypes) are classified by their specific pathogenic mechanisms such as toxins (ENTEROTOXIGENIC ESCHERICHIA COLI), etc. Alkalescens-Dispar Group,Bacillus coli,Bacterium coli,Bacterium coli commune,Diffusely Adherent Escherichia coli,E coli,EAggEC,Enteroaggregative Escherichia coli,Enterococcus coli,Diffusely Adherent E. coli,Enteroaggregative E. coli,Enteroinvasive E. coli,Enteroinvasive Escherichia coli
D005814 Genes, Viral The functional hereditary units of VIRUSES. Viral Genes,Gene, Viral,Viral Gene
D000251 Adenosine Triphosphatases A group of enzymes which catalyze the hydrolysis of ATP. The hydrolysis reaction is usually coupled with another function such as transporting Ca(2+) across a membrane. These enzymes may be dependent on Ca(2+), Mg(2+), anions, H+, or DNA. ATPases,Adenosinetriphosphatase,ATPase,ATPase, DNA-Dependent,Adenosine Triphosphatase,DNA-Dependent ATPase,DNA-Dependent Adenosinetriphosphatases,ATPase, DNA Dependent,Adenosinetriphosphatases, DNA-Dependent,DNA Dependent ATPase,DNA Dependent Adenosinetriphosphatases,Triphosphatase, Adenosine
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
D017122 Bacteriophage T4 Virulent bacteriophage and type species of the genus T4-like phages, in the family MYOVIRIDAE. It infects E. coli and is the best known of the T-even phages. Its virion contains linear double-stranded DNA, terminally redundant and circularly permuted. Bacteriophage T2,Coliphage T2,Coliphage T4,Enterobacteria phage T2,Enterobacteria phage T4,Phage T2,Phage T4,T2 Phage,T4 Phage,Phage, T2,Phage, T4,Phages, T2,Phages, T4,T2 Phages,T2, Enterobacteria phage,T4 Phages

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