Identification of a nucleic acid‐binding region within the largest subunit ofDrosophila melanogasterRNA polymerase II

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Abstract
The largest and the second‐largest subunit of the multisubunit eukaryotic RNA polymerases are involved in interaction with the DNA template and the nascent RNA chain. Using Southwestern DNA‐binding techniques and nitrocellulose filter binding assays of bacterially expressed fusion proteins, we have identified a region of the largest, 215‐kDa, subunit ofDrosophilaRNA polymerase II that has the potential to bind nucleic acids nonspecifically. This nucleic acid‐binding region is located between amino acid residues 309–384 and is highly conserved within the largest subunits of eukaryotic and bacterial RNA polymerases. A homology to a region of the DNA‐binding cleft ofEscherichia coliDNA polymerase I involved in binding of the newly synthesized DNA duplex provides indirect evidence that the nucleic acid‐binding region of the largest subunit participates in interaction with double‐stranded nucleic acids during transcription. The nonspecific DNA‐binding behavior of the region is similar to that observed for the native enzyme in nitrocellulose filter binding assays and that of the separated largest subunit in Southwestern assays. A high content of basic amino acid residues is consistent with the electrostatic nature of nonspecific DNA binding by RNA polymerases.
Funding Information
  • Deutsche Forschungsgemeinschaft (SFB 229)
  • EC (SCVCT 91/0702)