Cyclic adenosine‐5′‐trimetaphosphate phosphorylates a histidine residue nearby the initiating substrate binding site of Escherichia coli DNA‐dependent RNA‐polymerase

in the presence of a promoter template acts as an efficient affinity reagent towards the center of initiating substrate binding of DNA-dependent RNApolymerase of Escherichia coli. It was found that ATMP inactivates RNA-polymerase, the enzyme being protected from inactivation by ATP. On the contrary, addition of [w~‘P]UTP causes acceleration of the inactivation by a factor of 10-20. Radioactivity of [Q-~‘P] UTP under these conditions is bound covalently by the /3-subunit of the enzyme. These data suggested that the result of the affinity modification was phosphorylation of an ammo acid residue by an oligonucleotide synthesized by RNA-polymerase from ATMP and [a-32P]UTP. The present studies was aimed at the determination of the length of this covalently-bound oligonucleotide and at elucidation of the nature of the phosphorylated amino acid residue.