Elongation factor 1 (EF-1) consists of four subunits: the ± subunit catalyzes the GTP–dependent binding of aminoacyl-tRNA to ribosomes while the ², γ, and δ subunits catalyze GDP/GTP exchange on EF-1±. Phosphorylation of the β subunit of EF-1 from rabbit reticulocytes by casein kinase II was stimulated up to 22-fold by polylysine, while basic proteins or polyarginine enhanced phosphorylation to a lesser extent. When physiological components of protein synthesis were examined as potential modulators of phosphorylation, ribosomal subunits had no effect, tRNA and poly(U) inhibited the phosphotransferase reaction, and GDP stimulated the initial rate of phosphorylation of EF-1β up to 3.8-fold; the degree of stimulation could be correlated with the amount of α subunit present in EF-1. No stimulation was observed with other nucleotides. Phosphorylation of EF-1β was on serine, and two-dimensional phosphopeptide mapping showed a single tryptic phosphopeptide in the presence of GDP or polylysine; the peptide was identical to that obtained with EF-1 phosphorylated in reticulocytes incubated with [32P] orthophosphate. EF-1 δ was also phosphorylated by casein kinase II, but only in the presence of GDP. Kinetic data showed GDP stimulated phosphorylation by increasing the Vmax with both the β and δ subunits. The GDP-dependent stimulation of phosphorylation was specific for EF-1 and was not observed with calmodulin, βcasein B, or c-Myc. When the catalytic subunit of casein kinase II, cloned and expressed in Escherichia coli, was used, only EF-1β was phosphorylated; stimulation by GDP was approximately the same as that observed with the holoenzyme. Since guanine nucleotides bind only to EF-1α, GDP could regulate phosphorylation of EF-1β and δ only when present in the EF-1αβγδ complex. Thus, phosphorylation of EF-1β and δ by casein kinase II would occur primarily at a specific point in the elongation cycle, when EF-1α·GDP is associated with EF-1βγδ following the GTP-dependent binding of aminoacyl-tRNA to the ribosome.
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