Kinetics of biphasic reconstitution of tobacco mosaic virus in vitro.

The kinetics of the in vitro reconstitution of tobacco mosaic virus from its RNA and protein were studied by measuring the increase in turbidity, the development of ribonuclease-resistant infectivity, the emcapsidation of the terminal ends of the RNA, and the growth of rod length. The results showed that the reconstitution reaction consists of two processes in which the direction, timing, and rate of assembly are different. Rapid elongation of particles toward the 5' end of the RNA proceeds in the first 5-7 min to give intermediate particles of 260 nm in length in which only the 5' terminus of the RNA is encapsidated. The subsequent process requires 30-50 min, is accompanied by a slow increase in turbidity, and gives rise to rods of the full length, 300 nm. The 3' terminus becomes RNase resistant by this process with concomitant development of ribonuclease-resistant infectivity, showing that the 3'-distal portion of the RNA is encapsidated in the direction of 5' to 3'. The rate of rod elongation by the second process is less than 1/10 of that by the first process.