Process of biosynthetic protein folding determines the rapid formation of native structure

Abstract

Biosynthetic folding, beginning with the growing nascent chain and leading to the biologically active structure within its proper cellular context, is one function shared by all proteins. We show that the bacterial luciferase β subunit reaches its final native form in the αβ heterodimer much more rapidly during biosynthetic folding than during refolding from urea. The rate of formation of active enzyme is determined by a short-lived folding intermediate, which is able to associate with the α subunit very rapidly following release from the ribosome. This intermediate appears to involve a transient interaction of the C-terminal region of the β subunit, a region distant from the subunit interface, but intimately involved in heterodimerization. Refolding of the β subunit under similar conditions proceeds much more slowly. We have characterized both pathways and show that the basic difference between biosynthetic folding and refolding from urea is that the newly synthesized β subunit enters the folding pathway at a point beyond the slow, rate-determining step that limits the rate of the renaturation process and constitutes a kinetic trap. This mechanism embodies a major strategy, the avoidance of slow-folding intermediates and kinetic traps, that may be employed by many proteins to achieve fast and efficient biosynthetic folding.