Partial purification and characterization of a novel dna glycosylase from neisseria mucosa

Simon G. Nyaga, Robert (Stephen) Lloyd

Research output: Contribution to journalArticle

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Abstract

In an effort to identify functional, if not structural homologs of T4 endonuclease V, various strains of bacterial cell-free extracts were screened using site-specific pyrimidine dimer and 6-4 photoproduct containing duplex 49-mer. Several strains showed pyrimidine dimer nicking activity including Neisseria mucosa. Surprisingly, none of these strains showed 6-4 photoproduct nicking activity. We undertook the task of purifying the enzyme from Neisseria mucosa in order to obtain sufficient quantities for biochemical characterization. This enzyme has been partially purified by two Chromatographie steps: single stranded DNA agarose and heparin sepharose, and the products analyzed by silver staining. Polyclonal antibodies against endonuclease V did not cross-react with the Neisseria mucosa enzyme. Furthermore, the NH2-terminal sequence of the Neisseria mucosa protein did not show nomology with T4 endonuclease V although it showed high homology with a segment of Drosophila DNA replication licensing factor. A covalent imino enzyme-DNA intermediate was revealed by reduction with sodium borohydride (NaBH4). Thus, we have revealed a novel DNA glycosylase in Neisseria mucosa with qualitatively different pyrimidine dimer nicking activity and similar pyrimidine dimer incision mechanism compared to T4 endonuclease V.

Original languageEnglish (US)
JournalFASEB Journal
Volume10
Issue number6
StatePublished - 1996
Externally publishedYes

Fingerprint

Neisseria mucosa
glycosylases
Pyrimidine Dimers
pyrimidines
Purification
Enzymes
enzymes
agarose
Deoxyribonuclease (Pyrimidine Dimer)
DNA Glycosylases
single-stranded DNA
Silver Staining
Single-Stranded DNA
DNA
heparin
DNA replication
Licensure
Cell Extracts
polyclonal antibodies
DNA Replication

ASJC Scopus subject areas

  • Agricultural and Biological Sciences (miscellaneous)
  • Biochemistry, Genetics and Molecular Biology(all)
  • Biochemistry
  • Cell Biology

Cite this

Partial purification and characterization of a novel dna glycosylase from neisseria mucosa. / Nyaga, Simon G.; Lloyd, Robert (Stephen).

In: FASEB Journal, Vol. 10, No. 6, 1996.

Research output: Contribution to journalArticle

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