Hi. Peters et al., PHOSPHORYLATION OF ELONGATION-FACTOR 1(EF-1) BY PROTEIN-KINASE-C STIMULATES GDP GTP-EXCHANGE ACTIVITY/, European journal of biochemistry, 234(2), 1995, pp. 550-556
Phosphorylation of the alpha, beta and delta subunits of elongation fa
ctor (EF) 1 by protein kinase C results in stimulation of elongation a
ctivity up to threefold both in vivo and in vitro [Venema, R. C., Pete
rs, H. I. & Traugh, J. A. (1991) J. Biol. Chem. 266, 11993-11998; Vene
ma, R. C., Peters, H. I. & Traugh, J. A. (1991) J. Biol. Chem. 266, 12
574-12580]. The alpha subunit catalyzes the GTP-dependent binding of a
mino-acyl-tRNA to the ribosome, while the beta gamma and delta subunit
s of EF-1 catalyze exchange of the residual GDP on EF-1 alpha for GTP.
To determine whether the change in elongation rate following phosphor
ylation by protein kinase C is due to stimulation of GDP/GTP exchange
activity. EF-1 and EF-1 . valyl-tRNA-synthetase have been purified fro
m rabbit reticulocytes, phosphorylated in vitro by protein kinase C an
d the effect of phosphorylation on nucleotide-exchange activity analyz
ed. The alpha, beta and delta subunits are phosphorylated only on seri
ne, and phosphopeptide maps show distinct phosphopeptides for each sub
unit. Following quantitative phosphorylation of EF-1 by protein kinase
C on the alpha, beta and delta subunits, a twofold enhancement of the
rate of nucleotide exchange over the non-phosphorylated controls is o
bserved with EF-1 and EF-1 . valyl-tRNA synthetase. Stimulation of nuc
leotide exchange results in a two-fold increase in the formation of EF
-1 alpha . GTP . Phe-tRNA, leading to an increased rate of binding of
Phe-tRNA to ribosomes. The magnitude of stimulation of the exchange ra
te is similar to that reported previously for the rate of elongation f
ollowing phosphorylation of EF-1 by protein kinase C. Thus, the enhanc
ement of EF-1 activity in response to 4 beta-phorbol 12-myristate 13-a
cetate appears to be due to stimulation of the rate of GDP/GTP exchang
e following phosphorylation of EF-1 by protein kinase C.