The herpes simplex virus type 1 origin-binding protein carries out origin specific DNA unwinding and forms unwound stem-loop structures

Alexander M. Makhov, Paul E. Boehmer, I. Robert Lehman, Jack D. Griffith

Research output: Contribution to journalArticle

45 Scopus citations

Abstract

The UL9 protein of herpes simplex virus type 1 (HSV-1) binds specifically to the HSV-1 ori(S) and ori(L) origins of replication, and is a DNA helicase and DNA-dependent NTPase. In this study electron microscopy was used to investigate the binding of UL9 protein to DNA fragments containing ori(S). In the absence of ATP, UL9 protein was observed to bind specifically to ori(S) as a dimer or pair of dimers, which bent the DNA by 35° ± 15° and 86° ± 38° respectively, and the DNA was deduced to make a straight line path through the protein complex. In the presence of 4 mM ATP, binding at ori(S) was enhanced 2-fold, DNA loops or stem-loops were extruded from the UL9 protein complex at ori(S), and the DNA in them frequently appeared highly condensed into a tight rod. The stem-loops contained from a few hundred to over one thousand base pairs of DNA and in most, ori(S) was located at their apex, although in some, ori(S) was at a border. The DNA in the stem-loops could be stabilized by photocrosslinking, and when Escherchia coli SSB protein was added to the incubations, it bound the stem-loops strongly. Thus the DNA strands in the stem-loops exist in a partially paired, partially single-stranded state presumably making them available for ICP8 binding in vivo. These observations provide direct evidence for an origin specific unwinding by the HSV-1 UL9 protein and for the formation of a relatively stable four-stranded DNA in this process.

Original languageEnglish (US)
Pages (from-to)1742-1750
Number of pages9
JournalEMBO Journal
Volume15
Issue number7
StatePublished - Apr 1 1996

Keywords

  • Electron microscopy
  • Helicase
  • Herpes simplex virus
  • Origin
  • UL9 protein

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Biochemistry, Genetics and Molecular Biology(all)
  • Immunology and Microbiology(all)

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