Discovery and characterization of the cryptic psi subunit of the pseudomonad DNA replicase. Journal Article uri icon

Overview

abstract

  • We previously reconstituted a minimal DNA replicase from Pseudomonas aeruginosa consisting of alpha and epsilon (polymerase and editing nuclease), beta (processivity factor), and the essential tau, delta, and delta' components of the clamp loader complex (Jarvis, T., Beaudry, A., Bullard, J., Janjic, N., and McHenry, C. (2005) J. Biol. Chem. 280, 7890-7900). In Escherichia coli DNA polymerase III holoenzyme, chi and Psi are tightly associated clamp loader accessory subunits. The addition of E. coli chiPsi to the minimal P. aeruginosa replicase stimulated its activity, suggesting the existence of chi and Psi counterparts in P. aeruginosa. The P. aeruginosa chi subunit was recognizable from sequence similarity, but Psi was not. Here we report purification of an endogenous replication complex from P. aeruginosa. Identification of the components led to the discovery of the cryptic Psi subunit, encoded by holD. P. aeruginosa chi and Psi were co-expressed and purified as a 1:1 complex. P. aeruginosa chiPsi increased the specific activity of tau(3)deltadelta' 25-fold and enabled the holoenzyme to function under physiological salt conditions. A synergistic effect between chiPsi and single-stranded DNA binding protein was observed. Sequence similarity to P. aeruginosa Psi allowed us to identify Psi subunits from several other Pseudomonads and to predict probable translational start sites for this protein family. This represents the first identification of a highly divergent branch of the Psi family and confirms the existence of Psi in several organisms in which Psi was not identifiable based on sequence similarity alone.

publication date

  • December 1, 2005

has subject area

Date in CU Experts

  • October 1, 2013 12:09 PM

Full Author List

  • Jarvis TC; Beaudry AA; Bullard JM; Ochsner U; Dallmann HG; McHenry CS

author count

  • 6

citation count

  • 9

Other Profiles

International Standard Serial Number (ISSN)

  • 0021-9258

Electronic International Standard Serial Number (EISSN)

  • 1083-351X

Additional Document Info

start page

  • 40465

end page

  • 40473

volume

  • 280

issue

  • 49