Assembly of adenovirus type 2 fiber synthesized in cell-free translation system

Abstract

Physicochemical and functional analyses of the translation products of fiber mRNA in rabbit reticulocyte lysate suggested that fiber polypeptide chains (monomers) were capable of self-assembling in vitro, forming trimeric fibers (trimers) without direct intervention of any other adenovirus-coded protein or cell nuclear matrix component. Kinetic studies showed that trimer formation occurred at a rate six times lower than that of fiber polypeptide synthesis. Fiber assembly was found to be relatively inefficient in vitro, with only 25-30% fiber polypeptides trimerized after 4-h translation reaction. The rate constant for fiber subunit assembly, extrapolated from the kinetic curves of trimer formation, was found to be in the order of magnitude of 10(5) M-1 s-1, with a t 1/2 of 1.3 h at 30 degrees C. A latence phase of approximately 40 min in the appearance of the first detectable trimers indicated that fiber assembly did not occur co-translationally, suggesting the existence of rate-limiting intermediate step(s) during assembly.