Purification and properties of a high-molecular-mass complex between Val-tRNA synthetase and the heavy form of elongation factor 1 from mammalian cells.
In extracts of various mammalian tissues obtained in the presence of protease inhibitors Val-tRNA synthetase exists exclusively as a complex with a molecular mass of about 800 kDa. This complex was purified by gel filtration and two HPLC steps and contained five different polypeptides with molecular masses of 140, 50, 50, 40 and 30 kDa. The complex seems to have no tissue or species specificity, because preparations with identical polypeptide composition were obtained by the same method from rabbit liver and reticulocytes, and rat and beef liver. Four low-molecular-mass polypeptides were identified by two-dimensional electrophoresis as subunits of the heavy form of elongation factor 1 (EF-1H). The complex possesses the activity of EF-1 in the poly(U)-directed translation system, indicating that EF-1H is an integral part of the complex. Gel filtration of the tissue extracts reveals three different peaks of EF-1 activity, corresponding to EF-1 alpha, EF-1H and the high-molecular-mass complex of Val-tRNA synthetase and EF-1H. All activity of Val-tRNA synthetase and about 25% of EF-1 activity are associated with the complex. Different forms of EF-1 revealed no significant differences in the nucleotide-binding properties, but the complex of Val-tRNA synthetase with EF-1H was 10 times more active in the poly(U)-directed binding of Phe-tRNAPhe to ribosomes than EF-1H. These results strongly suggest that the complex of Val-tRNA synthetase with EF-1H is a novel functionally active individual form of EF-1.
A. N. Bakh Institute of Biochemistry, Academy of Sciences of USSR, Moscow.
Y A
YA
Motorin
A D
AD
Wolfson
D
D
Löhr
A F
AF
Orlovsky
K L
KL
Gladilin
D000818:Animals; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D008099:Liver / Q000201:enzymology; D008970:Molecular Weight; D020648:Peptide Elongation Factor 1; D010445:Peptide Elongation Factors / Q000302:isolation & purification* / Q000378:metabolism; D011072:Poly U / Q000378:metabolism; D012360:RNA, Transfer, Phe / Q000378:metabolism; D011817:Rabbits; D012156:Reticulocytes / Q000201:enzymology* / Q000378:metabolism; D012261:Ribonucleoproteins / Q000378:metabolism; D012270:Ribosomes / Q000201:enzymology / Q000378:metabolism; D014637:Valine-tRNA Ligase / Q000302:isolation & purification* / Q000378:metabolism
D020648:Peptide Elongation Factor 1; D010445:Peptide Elongation Factors; D012360:RNA, Transfer, Phe; D012261:Ribonucleoproteins; D011072:Poly U; D014637:Valine-tRNA Ligase