Demonstration of a guanosine triphosphate-dependent enzymatic binding of aminoacyl-ribonucleic acid to Escherichia coli ribosomes.

Abstract

Multiple enzyme fractionS< have been shown to be required for the synthesis of polypeptides from aminoacyl-sRNA in both mammalian and microbial systems.1'6 In the reticulocyte system two enzyme fractions have been founid to be required for the polyuridylic acid-(poly U)-directed synthesis of polyphenylalanine.7 One of these fractions was found to enihance the binding of phenylalanyl-sRNA to reticulocyte ribosomes in the presence of guanosine 5'-triphosphate (GTP) at concentrationis of Mlg++ that are close to the optimum for the over-all polymerization reaction.8' Three complemenitary polymerization factors isolated from extracts of Escherichia coli and Pseudomonas fluorescens10 have been shown to be required for polyphenylalanine synthesis with washed E. coli ribosomes. Several investigators"1-"3 have demonstrated that poly U-directed binding of phenylalanylsRNA to ribosomes occurs at high concentrations of i\Ig++ in the absence of GTP and soluble enzyme fractions (nonenzymatic binding); however, no enhancement of the biniding of phenylalanyl-sRNA to ribosomes in the presence of GTP and purified enzyme fractionis (enizymatic biniditng) has beeni observed previously in microbial systems.12 13 In the present study, ani extract of L'. coli W has been separated into two fractionls, both of which are required for the transfer of aminoacyl-sRNA to polypeptide with washed E. coli W ribosomes. One of these fractionis was founid to be required for the GTP-depeiidenit, enzymatic binding of aminoacyl-sRNA to E. coli ribosomes. These findings appear to be analogous to those previously reported in the reticulocyte system.8'