Polynucleotide phosphorylase in ribosomes from Escherichia coli

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Abstract
The stimulation of ribosome autodegradation into ribonucleoside 5[image]-phosphates by orthophosphate, has been examined by using inorganic [p32]phosphate and P32 -labelled ribonucleic acid. Dialysed preparations of disrupted Escherichia coli incorporated inorganic [P32]ortho-phosphate into the terminal phosphate of ribonucleoside 5[image]-diphosphates and also into an unidentified substance X. Arsenate did not alter the rate of degradation of RNA but reduced the extent of incorporation into nucleoside 5[image]-diphosphates and inhibited the formation of substance X. A ribosome preparation, incubated under similar conditions, also produced ribonucleoside 5[image]-diphosphates with P32 confined to the terminal phosphates of nucleoside 5[image]-diphosphates but not substance X. Dialysis reduced the rate of the process by about 50%. Arsenate did not affect the rate but reduced the concentrations of nucleoside 5[image]-diphosphates produced. An increase in the rate of the process, as the concentration of orthophosphate was increased, was accompanied by a progressive increase in the concentrations of nucleoside 5[image]-diphosphates in the products. The relative concentrations of cytidine 5[image]-diphosphate, adenosine 5[image]-diphosphate and uridine 5[image]-diphosphate produced in the presence of m[image]-20 m[image]-orthophosphate remained about constant; guanosine 5[image]-diphosphate, however, changed from the least prominent to the most prominent diphosphates as the concentration of orthophosphate was increased from m[image] to 2 m[image]. After the exhaustive auto-degradation of ribosomes and the addition of p32-labelled RNA, the specific activities of the nucleoside 5[image]-diphosphates exceeded those of the nucleoside 5[image] -monophosphates, suggesting that the nucleoside 5[image]-diphosphates are produced directly from RNA by phosphorolysis. The results suggest that both a polynucleotide phosphodiesterase and a polynucleotide phosphorylase are present in ribosomes from Escherichia coli and that the phosphorylase, during the early stages of the process, accounts for 14 or 36% of the total activity when 5 m[image]- or 20 m[image]-orthophosphate respectively is present.