The positively charged surface of herpes simplex virus UL42 mediates DNA binding

Gloria Komazin-Meredith, Webster L. Santos, David J. Filman, James M. Hogle, Gregory L. Verdine, Donald M. Coen

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

Herpes simplex virus DNA polymerase is a heterodimer composed of UL30, a catalytic subunit, and UL42, a processivity subunit. Mutations that decrease DNA binding by UL42 decrease long chain DNA synthesis by the polymerase. The crystal structure of UL42 bound to the C terminus of UL30 revealed an extensive positively charged surface ("back face"). We tested two hypotheses, 1) the C terminus of UL30 affects DNA binding and 2) the positively charged back face mediates DNA binding. Addressing the first hypothesis, we found that the presence of a peptide corresponding to the UL30 C terminus did not result in altered binding of UL42 to DNA. Addressing the second hypothesis, previous work showed that substitution of four conserved arginine residues on the basic face with alanines resulted in decreased DNA affinity. We tested the affinities for DNA and the stimulation of long chain DNA synthesis of mutants in which the four conserved arginine residues were substituted individually or together with lysines and also a mutant in which a conserved glutamine residue was substituted with an arginine to increase positive charge on the back face. We also engineered cysteines onto this surface to permit disulfide cross-linking studies. Last, we assayed the effects of ionic strength on DNA binding by UL42 to estimate the number of ions released upon binding. Our results taken together strongly suggest that the basic back face of UL42 contacts DNA and that positive charge on this surface is important for this interaction.

Original languageEnglish (US)
Pages (from-to)6154-6161
Number of pages8
JournalJournal of Biological Chemistry
Volume283
Issue number10
DOIs
StatePublished - Mar 7 2008

Fingerprint

Simplexvirus
Viruses
DNA
Arginine
DNA-Directed DNA Polymerase
Glutamine
Disulfides
Alanine
Osmolar Concentration
Ionic strength
Lysine
Cysteine
Catalytic Domain
Substitution reactions
Ions
Crystal structure
Peptides
Mutation

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Cell Biology
  • Molecular Biology

Cite this

Komazin-Meredith, Gloria ; Santos, Webster L. ; Filman, David J. ; Hogle, James M. ; Verdine, Gregory L. ; Coen, Donald M. / The positively charged surface of herpes simplex virus UL42 mediates DNA binding. In: Journal of Biological Chemistry. 2008 ; Vol. 283, No. 10. pp. 6154-6161.
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The positively charged surface of herpes simplex virus UL42 mediates DNA binding. / Komazin-Meredith, Gloria; Santos, Webster L.; Filman, David J.; Hogle, James M.; Verdine, Gregory L.; Coen, Donald M.

In: Journal of Biological Chemistry, Vol. 283, No. 10, 07.03.2008, p. 6154-6161.

Research output: Contribution to journalArticle

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