Purification of chloroplast elongation factors

Abstract

This chapter describes the procedures for the purification of elongation factor Tu (EF-Tu) and elongation factor G (EF-G) from isolated spinach chloroplasts and of ET-Tu starting from the intact cells of Euglena gracilis . Elongation factors are soluble proteins that participate in protein synthesis and in the elongation cycle. Chloroplasts contain elongation factors structurally and functionally analogous to those present in bacteria. In bacteria, three factors are isolated and characterized: EF-G, EF-Tu, and elongation factor Ts (EF-Ts). EF-G is responsible for the translocation of peptidyl-tRNA on the ribosome, whereas EF-Tu binds and carries to the ribosome the different aminoacyl-tRNAs. EF-Ts, involved in the detachement of Guanosine diphosphate (GDP) from the EF-Tu GDP complex, may not be required in vitro , whereas it is presumably necessary in vivo where it may exist in a stoichiometric complex with EF-Tu to give elongation factor T (EF-T). The assay systems for chloroplast elongation factors rely on the functional interchangeability of ribosomes and elongation factors from these organelles and those from Escherichia coli . The chloroplast elongation factors are proteins different from those present in the cell cytoplasm and those present in mitochondria.