Deoxyribo ucleic acid-directed synthesis of ribonucleic acid by an enzyme from Escherichia coli.

l’rotein structure is under genetic control;’-3 yet the precise mechanism by which DNA$ influences the formation of specific amino acid sequences in proteins is unknown. Several years ago, it was discovered that infection of Escherichia coli with certain virulent bacteriophages induces the formation of an RNA fraction possessing both a high metabolic turnover rate and a base composition corresponding to the DNA of the infecting v i r ~ s . ~ ~ The existence of an analogous R E A component in noninfected cells has also been demonstrated; in this instance, however, the base composition of the RNA resembles that of the cellular DNA.’. These observations focused attention on the possible role of this type of RNA in protein synthesis, and some of the evidence consistent with this view has recently been surnmari~ed.~ Until recently there was no known enzymatic mechanism for a DNA-directed synthesis of RNA. Polynucleotide phosphorylasel0, l 1 although it catalyzes the synthesis of polyribonucleotides, does not by itself provide a mechanism for the formation of RNA with a specific sequence of nucleotides. The one instance in which a unique sequence of nucleotides is produced involves the limited addition of nucleotides exclusively to the end of pre-existing polynucleotide chains. 1 2 1 4 Our efforts were therefore directed toward examining alternate mechanisms for RNA synthesis, and in particular one in which DNA might dictate the nucleotide sequence of the RNA. In the present paper, we wish to report the isolation and some properties of an RNA polymerase from E. coli which, in the presence of DNA and the four naturally occurring ribonucleoside triphosphates, produces RNA with a base composition complementary to that of the DNA. Within the last year, several laboratories have reported similar findings with enzyme preparations from bacterial as well as from plant and animal source^.^^-^^ I n the following paper, the effect of enzymatically synthesized RNA on the rate and extent of amino acid incorporation into protein by E. coli ribosomes in the presence of a soluble protein fraction is described. Unlabeled ribonucleoside diand triphosphates were purchased from the Sigma Biochemical Corporation and the California Corporation for Biochemical Resenrch. 8-C14-1abeled ATP was purchased from the Schwartz Biochemical Company; the other, uniformly labeled, C1* ribonucleoside triphosphates were prepared enzymatically from the corresponding monophosphate derivatives55 isolated from the RN.4 of Chromatium grown on CL402 as sole carbon source.*6 CTP labeled with P31 in the eater phosphate was obtained by enzymatic phosphorylation of CMPS* prepared according to Hurwitz.27 The deoxyribonucleoside triphosphates were obtained by the procedure of Lehman et al.25 DNA from Aerobacter aerogenes, Mycobacterium phlei, and bacteriophages T2, T5, T6 was prepared as described previously.29 Unlabeled and P32 labeled DNA from E. coli were prepared as previously de~cr ibed .~’ d-AT and d-GC polymers were prepared according to Schachman et al.32 and Radding et respectively. Transforming DNA from Ban’ZZus subliZisJ4 was a gift from E. %’. Nester, and DNA from phage 0X