Mapping of trypsin cleavage and antibody-binding sites and delineation of a dispensable domain in the β subunit of escherichia coli RNA polymerase

S. Borukhov, K. Severinov, M. Kashlev, A. Lebedev, I. Bass, G. C. Rowland, P. P. Lim, R. E. Glass, V. Nikiforov, A. Goldfarb

Research output: Contribution to journalArticlepeer-review

50 Scopus citations

Abstract

We have mapped principal sites in the Escherichia coli RNA polymerase molecule that are exposed to attack by trypsin under limited proteolysis conditions. The 1342-amino acid-long β subunit is alternatively cleaved at Arg903 or Lys909. The cleavage occurs adjacent to a dispensable domain (residues 940-1040) that is absent in the homologous RNA polymerase subunits from chloroplasts, eukaryotes, and archaebacteria. In E. coli, this region can be disrupted with genetic deletions and insertions without the loss of RNA polymerase function. Insertion of 127 amino acids into this region introduces a new highly labile site for trypsin proteolysis. The dispensable domain carries the epitope for monoclonal antibody PYN-6 (near residue 1000), which can be used for anchoring the catalytically active enzyme on a solid support. We also report the identification of a secondary trypsin cleavage at Arg81 of the β' subunit within a putative zinc-binding domain that is conserved in prokaryotes and chloroplasts.

Original languageEnglish (US)
Pages (from-to)23921-23926
Number of pages6
JournalJournal of Biological Chemistry
Volume266
Issue number35
StatePublished - 1991
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Fingerprint

Dive into the research topics of 'Mapping of trypsin cleavage and antibody-binding sites and delineation of a dispensable domain in the β subunit of escherichia coli RNA polymerase'. Together they form a unique fingerprint.

Cite this