Phosphorylation of glycogen synthase and of the beta subunit of eukaryotic initiation factor two by a common protein kinase.

A A Depaoli-Roach,P J Roach,K Pham,G Kramer,B Hardesty

doi:10.1016/s0021-9258(19)52475-9

Abstract

A protein kinase from rabbit reticulocytes, able to phosphorylate the beta subunit of eukaryotic initiation factor 2 (eIF-2), has been demonstrated to phosphorylate also glycogen synthase. A glycogen synthase kinase (PC0.7) from rabbit skeletal muscle has been shown to phosphorylate the beta subunit of eIF-2. Comparison of highly purified preparations of the two protein kinases has indicated several similarities of properties. 1) Both enzymes were associated with two major polypeptide species, alpha (Mr = 43,000) and beta (Mr = 25,000), and exhibited apparent native molecular weights of 176,000-180,000 by gel filtration and 130,000-140,000 by sucrose density gradient sedimentation. 2) Both enzymes phosphorylated glycogen synthase, eIF-2 beta, phosvitin, and casein and were effective in utilizing GTP and ATP as phosphoryl donors. 3) Both enzymes displayed the same chromatographic behavior on phosvitin-Sepharose, phosphocellulose, and DEAE-cellulose. 4) Both enzymes underwent an autophosphorylation of the beta polypeptide when incubated with ATP and Mg2+. On the basis of these and other properties, we propose that the two protein kinases, if not identical, are very similar enzymes.

Highlights

IntroductionWe conclude that the reticulocyte eIF-2Pkinase and the muscle PC"., are, if not identical, closely related protein kinases
On the basis of these and other properties, we propose that the two protein kinases, ifnot identical, tRNA to 40 S ribosomal subunits due to eIF-2 was determined by the retention of 13'S]Met-tRNAr . 40 S complex on Millipore fdters, as reported previously [16]
Protein Substrates-Glycogen synthase was purified from rabbit skeletal muscle as described previously and had properties similar to those reported [17, 18].eIF-2 was purified from the reticulocyte 0.5 M KC1-ribosomal wash fraction by chromatography on DEAE-cellulose and phosphocellulose as previously described [16].Phosvitin was obtained from Sigma

Summary

Introduction

We conclude that the reticulocyte eIF-2Pkinase and the muscle PC"., are, if not identical, closely related protein kinases. On the basis of these and other properties, we propose that the two protein kinases, ifnot identical, tRNA to 40 S ribosomal subunits due to eIF-2 was determined by the retention of 13'S]Met-tRNAr . Protein Substrates-Glycogen synthase was purified from rabbit skeletal muscle as described previously and had properties similar to those reported [17, 18].eIF-2 was purified from the reticulocyte 0.5 M KC1-ribosomal wash fraction by chromatography on DEAE-cellulose and phosphocellulose as previously described [16].Phosvitin was obtained from Sigma. Protein KinasePurification-Full details of the purification close to apparent homogeneity of the rabbit reticulocyte eIF-2P kinase are very similar enzymes. The final three purification steps were identical for both preparations and, briefly, involved binding to phosvitin-Sepharose, gel filtration on Bio-

Methods

Results

Conclusion