Mechanism of self-assembly of tobacco mosaic virus protein. I. Nucleation-controlled kinetics of polymerization.

The polymerization of tobacco mosaic virus protein has been found to proceed through metastable states under conditions where initially one of the two polymerization-linked protons is bound. These metastable polymers have been characterized and are found to be helical rods, which resemble the structure of equilibrium helical rods that form when both polymerization-linked protons are bound. At pH 6.5 and 20 °C the true equilibrium distribution of these helical rods has been shown to consist of sedimenting species that are much smaller, 24 to 34 S, than described previously, 100 to 200 S. The larger, non-equilibrium rods are produced by an overshoot in polymerization that results from the slow formation of 20 S nuclei followed by a very rapid elongation reaction. Generally, this sequence of rate processes is sensitive to the rate at which a reaction is initiated. In the present case it is the rate of heating or the rate of change of the pH that determines the reaction path and therefore the rate of attainment of equilibrium. In addition to the formation of metastable helical rods during polymerization overshoot, metastable 20 S aggregates can form when either equilibrium or non-equilibrium helical rods are depolymerized by cooling to 5 to 7 °C at pH 6.5. These 20 S aggregates are presumably two-turn disks or helices and can serve as nuclei for helical rod formation in subsequent polymerization reactions. Both helical rod and 20 S metastability are extremely sensitive to pH but, under carefully controlled conditions, the metastability is quite reproducible and reproducible nucleation-controlled polymerization kinetics can be observed even when polymerization-depolymerization cycling is carried out between branches of a hysteresis loop. Temperature- or pH-induced polymerization of tobacco mosaic virus protein can be made to proceed by the slow formation of 20 S, two-turn helix, nuclei followed by the rapid addition of one or more species comprising the 4 S protein. These results confirm a previously proposed kinetic mechanism for the non-equilibrium polymerization reaction (Scheele &; Schuster, 1974).