Mechanism of Bacteriophage T4 DNA Packaging ATPase

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Mechanism of Bacteriophage T4 DNA Packaging ATPase

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Title: Mechanism of Bacteriophage T4 DNA Packaging ATPase
Author: Gao, Song
Abstract: The DNA packaging motors of bacteriophages are the most powerful force–generating biological machines reported to date. The T4 DNA packaging motor is composed of the dodecameric portal vortex protein gp20 (61 kDa) and a pentamer of the large terminase protein gp17 (70 kDa). gp17 is a weak ATPase whose activity is dramatically stimulated either by the small terminase gp16 (18 kDa) or by assembly into a packaging motor. This ATPase activity, which is mapped to the N–terminal domain of gp17 is central to the packaging mechanism and energetically coupled to DNA translocation. In this study, the small and large terminase proteins from bacteriophages RB49 and KVP40, and small terminases from other T4 family phages were studied for ATPase activity and stimulation. The nuclease and translocase activities, which are also essential for DNA packaging, were as well tested. The results show that the interactions between gp16 and gp17 are highly specific. gp17 from the T4 phage is stimulated by its own gp16 but not by a gp16 from another family member phage. Extensive domain swapping experiments to switch the specificity of one phage terminase to another showed, surprisingly, that the interaction sites are localized to both the N– and C–terminal domains of gp16. But the most stringent specificity region was mapped to the C–terminal ∼20 amino acids. For gp17, partial change in specificity was observed when the N–terminal ∼80 amino acids were swapped. Moreover, M13 phage display library screening of peptides binding to T4 gp16 revealed several potential gp16 interacting sites on gp17, including a region close to the important ATPase coupling motif (TTT<sub>285–287</sub>). Combining these results with the available structural and modeling data on gp17 and gp16, a novel model involving multiple interaction sites on both gp17 and gp16 is proposed for the mechanism of ATPase stimulation. Multiple weak interactions between binding partners, one a molecular motor (gp17) and the other a regulator (gp16), might provide the greatest flexibility to fine–tune the functions of a complex motor: assembly, ATPase, DNA translocation and headful nuclease.
Description: Degree awarded: Ph.D. Biology. The Catholic University of America
URI: http://hdl.handle.net/1961/9190
Date: 2011-02-24


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