Ultrastable pRNA hexameric ring gearing hexameric phi29 DNA-packaging motor by revolving without rotating and coiling.
ABSTRACT Biomotors have previously been classified into two categories: linear and rotational motors. It has long been popularly believed that viral DNA packaging motors are rotation motors. We have recently found that the DNA-packaging motor of bacteriophage phi29 uses a third mechanism: revolution without rotation. phi29 motor consists of three-coaxial rings of hexameric RNA, a hexameric ATPase, and a dodecameric channel. The motor uses six ATP to revolve one helical turn of dsDNA around the hexameric ring of ATPase gp16. Each dodecameric segment tilts at a 30°-angle and runs anti-parallel to the dsDNA helix to facilitate translation in one direction. The negatively charged phosphate backbone interacts with four positively charged lysine rings, resulting in four steps of transition. This review will discuss how the novel pRNA meets motor requirements for translocation concerning structure, stoichiometry, and thermostability; how pRNA studies have led to the generation of the concept of RNA nanotechnology; and how pRNA is fabricated into nanoparticles to deliver siRNA, miRNA, and ribozymes to cancer and virus-infected cells.
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ABSTRACT: The DNA packaging motor of the bacteriophage ϕ29, comprising head-tail connector, ATPase, and pRNA, transports the viral DNA inside the procapsid against pressure differences of up to ∼60 atm during replication. Several models for the DNA packaging mechanism have been proposed, which attribute different roles to the connector, and require specific mechanical properties of the connector. To characterize these properties at the atomic level, and to understand how the connector withstands this large pressure, we have carried out molecular dynamics simulations of the whole connector both in equilibrium and under mechanical stress. The simulations revealed a quite heterogeneous distribution of stiff and soft regions, resembling that of typical composite materials that are also optimized to resist mechanical stress. In particular, the conserved middle α-helical region is found to be remarkably stiff, similar only to structural proteins forming viral shell, silk, or collagen. In contrast, large parts of the peripheral interface to the ϕ29 procapsid turned out to be rather soft. Force probe and umbrella sampling simulations showed that large connector deformations are remarkably reversible, and served to calculate the free energies required for these deformations. In particular, for an untwisting deformation by 12°, as postulated by the untwist-twist model, more than four times’ larger energy is required than is available from hydrolysis of one ATP molecule. Combined with previous experiments, this result is incompatible with the untwist-twist model. In contrast, our simulations support the recently proposed one-way revolution model and suggest in structural terms how the connector blocks DNA leakage. In particular, conserved loops at the rim of the central channel, which are in direct contact with the DNA, are found to be rather flexible and tightly anchored to the rigid central region. These findings suggest a check-valve mechanism, with the flexible loops obstructing the channel by interacting with the viral DNA.Biophysical Journal 03/2014; 106(6):1338-1348. DOI:10.1016/j.bpj.2014.01.028 · 3.97 Impact Factor
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ABSTRACT: Nanobiomotors perform various important functions in the cell, and they also emerge as potential vehicle for drug delivery. These proteins employ conserved ATPase domains to convert chemical energy to mechanical work and motion. Several archaeal nucleic acid nanobiomotors, such as DNA helicases that unwind double-stranded DNA molecules during DNA damage repair, have been characterized in details. XPB, XPD and Hjm are SF2 family helicases, each of which employs two ATPase domains for ATP binding and hydrolysis to drive DNA unwinding. They also carry additional specific domains for substrate binding and regulation. Another helicase, HerA, forms a hexameric ring that may act as a DNA-pumping enzyme at the end processing of double-stranded DNA breaks. Common for all these nanobiomotors is that they contain ATPase domain that adopts RecA fold structure. This structure is characteristic for RecA/RadA family proteins and has been studied in great details. Here we review the structural analyses of these archaeal nucleic acid biomotors and the molecular mechanisms of how ATP binding and hydrolysis promote the conformation change that drives mechanical motion. The application potential of archaeal nanobiomotors in drug delivery has been discussed.06/2014; 4:32. DOI:10.1186/2045-3701-4-32
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ABSTRACT: DNA damage attacks on bacterial cells have been known to activate the SOS response, a transcriptional response affecting chromosome replication, DNA recombination and repair, cell division and prophage induction. All these functions require double-stranded (ds) DNA translocation by ASCE hexameric motors. This review seeks to delineate the structural and functional characteristics of the SOS response and the SOS-regulated DNA translocases FtsK and RuvB with the phi29 bacteriophage packaging motor gp16 ATPase as a prototype to study bacterial motors. While gp16 ATPase, cellular FtsK and RuvB are similarly comprised of hexameric rings encircling dsDNA and functioning as ATP-driven DNA translocases, they utilize different mechanisms to accomplish separate functions, suggesting a convergent evolution of these motors. The gp16 ATPase and FtsK use a novel revolution mechanism, generating a power stroke between subunits through an entropy-DNA affinity switch and pushing dsDNA inward without rotation of DNA and the motor, whereas RuvB seems to employ a rotation mechanism that remains to be further characterized. While FtsK and RuvB perform essential tasks during the SOS response, their roles may be far more significant as SOS response is involved in antibiotic-inducible bacterial vesiculation and biofilm formation as well as the perspective of the bacteria-cancer evolutionary interaction.06/2014; 4:31. DOI:10.1186/2045-3701-4-31