Application of electrospray ionization mass spectrometry to study the hydrophobic interaction between the epsilon and theta subunits of DNA polymerase III

Protein Sci. 2004 Nov;13(11):2878-87. doi: 10.1110/ps.04889604. Epub 2004 Sep 30.


The interactions between the N-terminal domain of the epsilon (epsilon186) and theta subunits of DNA polymerase III of Escherichia coli were investigated using electrospray ionization mass spectrometry. The epsilon186-theta complex was stable in 9 M ammonium actetate (pH 8), suggesting that hydrophobic interactions have a predominant contribution to the stability of the complex. Addition of primary alkanols to epsilon186-theta in 0.1 M ammonium acetate (pH 8), led to dissociation of the complex, as observed in the mass spectrometer. The concentrations of methanol, ethanol, and 1-propanol required to dissociate 50% of the complex were 8.9 M, 4.8 M, and 1.7 M, respectively. Closer scrutiny of the effect of alkanols on epsilon186, theta, and epsilon186-theta showed that epsilon186 formed soluble aggregates prior to precipitation, and that the association of epsilon186 with theta stabilized epsilon186. In-source collision-induced dissociation experiments and other results suggested that the epsilon186-theta complex dissociated in the mass spectrometer, and that the stability (with respect to dissociation) of the complex in vacuo was dependent on the solution from which it was sampled.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • DNA Polymerase III / chemistry*
  • Enzyme Stability
  • Escherichia coli Proteins / chemistry
  • Hydrophobic and Hydrophilic Interactions
  • Protein Structure, Quaternary
  • Protein Subunits / chemistry
  • Spectrometry, Mass, Electrospray Ionization / methods*
  • Static Electricity


  • Escherichia coli Proteins
  • Protein Subunits
  • DNA Polymerase III