11 Regulation of In Vitro Protein Synthesis by Bacteriophage RNA by RNA Tertiary Structure

Abstract

INTRODUCTION
One of the central goals in the study of RNA phages is the elucidation of the regulation of protein synthesis in the infected cell. This has turned out to be a rather difficult task, and most studies of regulation of f2 protein synthesis involve studies of synthesis of the three phage proteins in cell-free extracts, using added phage RNA as template. It must be emphasized at the outset that there are considerable—and at present often insurmountable—obstacles in extrapolating such in vitro experiments to the case of regulation in the infected cell.

First, in even the most efficient bacterial cell-free systems available (preincubated, crude extracts of E. coli) (Webster et al. 1967), the amount of protein produced, expressed as micromoles of coat protein synthesized per micromole of RNA added, is about four, whereas in the infected cell this number is probably at least 10- to 40-fold greater (calculated from Fromageot and Zinder 1968; Nathans et al. 1969). Further, at best only about 20% of the f2 RNA added to cell lysates is ever attached to ribosomes (and presumably directing synthesis of proteins), and one can never be sure that the general properties of the added mRNA population are representative of the active fraction (Webster and Zinder 1967; H. Lodish, unpublished data). In most fractionated cell-free systems, containing washed ribosomes, purified initiation factors, and so forth, the efficiency of protein synthesis is much less. Clearly some factors or components are limiting in these systems, relative to the whole cell,...