Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.6.3.14 (ATP synthase)
 7,042 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We characterized nine helicase-deficient mutants of bacteriophage T7 helicase-primase protein (4A') prepared by random mutagenesis as reported in the accompanying paper (Rosenberg, A. H., Griffin, K., Washington, M. T., Patel, S. S., and Studier, F. W. (1996) J. Biol. Chem. 271, 26819-26824). Mutants were selected from each of the helicase-conserved motifs for detailed analysis to understand better their function. In agreement with the in vivo results, the mutants were defective in helicase activity but were active in primase function. dTTP hydrolysis, DNA binding, and hexamer formation were examined. Three classes of defective mutants were observed. Group A mutants (E348K, D424N, and S496F), defective in dTTP hydrolysis, lie in motifs 1a, 2, and 4 and are possibly involved in NTP binding/hydrolysis. Group B mutants (R487C and G488D), defective in DNA binding, lie in motif 4 and are responsible directly or indirectly for DNA binding. Group C mutants (G116D, A257T, S345F, and G451E) were not defective in any of the activities except the helicase function. These mutants, scattered throughout the protein, appear defective in coupling dTTPase activity to helicase function. Secondary structural predictions of 4A' and DnaB helicases resemble the known structures of RecA and F1-ATPase enzymes. Alignment shows a striking correlation in the positions of the amino acids that interact with NTP and DNA.
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PMID:Biochemical analysis of mutant T7 primase/helicase proteins defective in DNA binding, nucleotide hydrolysis, and the coupling of hydrolysis with DNA unwinding. 890 Jan 64

Bacteriophage T7 DNA helicase is a ring-shaped hexamer that catalyzes duplex DNA unwinding using dTTP hydrolysis as an energy source. Of the six potential nucleotide binding sites on the hexamer, we have found that three are noncatalytic sites and three are catalytic sites. The noncatalytic sites bind nucleotides with a high affinity, but dTTPs bound to these sites do not dissociate or hydrolyze through many dTTPase turnovers at the catalytic sites. The catalytic sites show strong cooperativity which leads to sequential binding and hydrolysis of dTTP. The elucidated dTTPase mechanism of the catalytic sites of T7 helicase is remarkably similar to the binding change mechanism of the ATP synthase. Based on the similarity, a general mechanism for hexameric helicases is proposed. In this mechanism, an F1-ATPase-like rotational movement around the single-stranded DNA, which is bound through the central hole of the hexamer, is proposed to lead to unidirectional translocation along single-stranded DNA and duplex DNA unwinding.
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PMID:The dTTPase mechanism of T7 DNA helicase resembles the binding change mechanism of the F1-ATPase. 914 81