T7 DNA Helicase
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T7 DNA Helicase is a hexameric motor protein that uses energy from dTTP hydrolysis to process unidirectionally along single stranded DNA, separating the two strands as progresses.
Crystal Structure
The crystal structure was solved to 3.0 Å resolution in 2000, Singleton MR, Sawaya MR, Ellenberger T, Wigley DB, ["Crystal Structure of T7 Gene 4 Ring Helicase Indicates a Mechanism for Sequential Hydrolysis of Nucleotides"], Cell. (2000) Jun 9; 101 (6): 589-600. as shown in the figure. In (A), notice that the separate subunits appear to be anchored through interactions between an alpha helix and an adjacent subunit. In (B), there are six sets of three loops. The red loop, known as loop II, contains three lysine residues and is thought to be involved in binding the ssDNA that is fed through the center of the enzyme.Mechanism of Sequential dTTP Hydrolysis
Crampton et al. have proposed a mechanism for the ssDNA-dependent hydrolysis of dTTP by T7 DNA Helicase as shown in the figure below. Crampton DJ, Mukherjee S, Richardson CC, ["DNA-Induced Switch from Independent to Sequential dTTP Hydrolysis in the Bacteriophage T7 DNA Helicase"], Molecular Cell. (2006) Jan 20; 21(2): 165-174. In their model, protein loops located on each hexameric subunit, each of which contain three lysine residues, sequentially interact with the negatively charged phosphate backbone of ssDNA. This interaction presumably causes a conformational change in the actively bound subunit, providing for the efficient release of dTDP from its dTTP binding site. In the process of dTDP release, the ssDNA is transferred to the neighboring subunit, which undergoes a similar process. Previous studies have already suggested that ssDNA is able to bind to two hexameric subunits simultaneously. Yu X, Hingorani MM, Patel SS, Egelman EH, ["DNA is bound within the central hole to one or two of the six subunits of the T7 DNA Helicase"], Nature Structural Biology. (1996) Sep; 3 (9): 740-3.
References
See also
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