Mechanism of One-Way Traffic of Hexameric Phi29 DNA Packaging Motor with Four Electropositive Relaying Layers Facilitating Antiparallel Revolution

Abstract

The importance of nanomotors in nanotechnology is akin to that of mechanical engines to daily life. The AAA+ superfamily is a class of nanomotors performing various functions. Their hexagonal arrangement facilitates bottom-up assembly for stable structures. The bacteriophage phi29 DNA translocation motor contains three coaxial rings: a dodecamer channel, a hexameric ATPase ring, and a hexameric pRNA ring. The viral DNA packaging motor has been believed to be a rotational machine. However, we discovered a revolution mechanism without rotation. By analogy, the earth revolves around the sun while rotating on its own axis. One-way traffic of dsDNA translocation is facilitated by five factors: (1) ATPase changes its conformation to revolve dsDNA within a hexameric channel in one direction; (2) the 30° tilt of the channel subunits causes an antiparallel arrangement between two helices of dsDNA and channel wall to advance one-way translocation; (3) unidirectional flow property of the internal channel loops serves as a ratchet valve to prevent reversal; (4) 5′–3′ single-direction movement of one DNA strand along the channel wall ensures single direction; and (5) four electropositive layers interact with one strand of the electronegative dsDNA phosphate backbone, resulting in four relaying transitional pauses during translocation. The discovery of a riding system along one strand provides a motion nanosystem for cargo transportation and a tool for studying force generation without coiling, friction, and torque. The revolution of dsDNA among 12 subunits offers a series of recognition sites on the DNA backbone to provide additional spatial variables for nucleotide discrimination for sensing applications.

Biological Nanomotors with a Revolution, Linear, or Rotation Motion Mechanism

“…A counterclockwise motion of the dsDNA in the T7 motor was observed when viewing from the N terminus of T7 connector (131) (Fig. 5B), in agreement with the direction of dsDNA revolution in the phi29 dsDNA-packaging motor (12,13,60,132) (Fig. 5A).…”

“…Based on their motion mechanisms, the biomotors are categorized into three classes: linear, rotary, and revolutional ( Fig. 1) (8)(9)(10)(11)(12)(13)(14)(15)(16). Rotation is the circular movement of an object around its own axis, resembling the Earth rotating on its axis in a complete cycle every 24 h. Revolution is the turning of an object around a second object, resembling the action of the Earth revolving around the sun one circle per year (Fig.…”

“…DNA was found to twist by as little as 1.5 degrees per base pair translocated, confirming a nonrotation mechanism (6), since 1.5°/bp ϫ 10.5 bp/helical turn ϭ 15.7°is far below the 360°per complete helical turn. This puzzle of "rotation motors that do not rotate" was not solved until the breakthrough of the discovery of revolution motion in 2013 by Guo's group (12)(13)(14)(15)(16) (Fig. 5).…”

“…14A) is related to the four relaying lysine layers embedded inside the phi29 connector channel inner wall ( Fig. 14B and C) (see below) (13,117,129).…”

“…The motor channels (connectors) of SPP1 (61), T7 (62), HK97, P22 (64), and phi29 (52) all adopt an antichiral arrangement between the left-handed motor connector subunits and the right-handed DNA helices during packaging ( Fig. 15) (12,13). No apparent homology among these portal proteins has been shown by sequence alignment.…”