Purification of Mitochondrial Uracil-DNA Glycosylase Using Ugi-Sepharose Affinity Chromatography

Bennett, Samuel; Shroyer, Mary Jane N.; Sung, Jung‐Suk; Mosbaugh, Dale W.

doi:10.1385/1-59259-284-8:211

Cited by 1 publication

(2 citation statements)

References 9 publications

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“…Ugi-Sepharose was prepared and affinity chromatography performed as described (24). Ugi-Sepharose was poured into a Micro Bio-Spin (Bio-Rad) column (250-l bed volume) and equilibrated in buffer B (30 mM Tris⅐HCl, pH 7.4͞1 mM EDTA͞1 mM DTT͞5% wt/vol glycerol).…”

Section: Methodsmentioning

confidence: 99%

“…This matrix contained covalently bound Ugi-protein that acts as a ligand to bind specifically and tightly Ung-family enzymes (24). It was anticipated that if the Ugi-sensitive uracil-DNA glycosylase activity detected in the Detection of uracil-DNA glycosylase activity by using a uracilcontaining oligodeoxynucleotide substrate.…”

Section: Detection Of Uracil-dna Glycosylase Activity By Using a Uracil-mentioning

confidence: 99%

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Escherichia coli nucleoside diphosphate kinase does not act as a uracil-processing DNA repair nuclease

Bennett

Chen²,

Mosbaugh

2004

Proc. Natl. Acad. Sci. U.S.A.

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Escherichia coli nucleoside diphosphate kinase (Ndk) catalyzes ATP-dependent synthesis of ribo-and deoxyribonucleoside triphosphates from the cognate diphosphate precursor. Recently, the Ndk polypeptide was reported to be a multifunctional base excision repair nuclease that processed uracil residues in DNA by acting sequentially as a uracil-DNA glycosylase inhibitor protein ( . USA 100, 13247-13252]. Here we demonstrate that the E. coli Ndk polypeptide lacked detectable uracil-DNA glycosylase activity and, hence, was incapable of acting as a uracil-processing DNA repair nuclease. This finding was based on the following observations: (i) uracil-DNA glycosylase activity did not copurify with Ndk activity; (ii) Ndk purified from E. coli ung ؊ cells showed no detectable uracil-DNA glycosylase activity; and (iii) Ndk failed to bind to a Ugi-Sepharose affinity column that tightly bound E. coli uracil-DNA glycosylase (Ung). Collectively, these observations demonstrate that the E. coli Ndk polypeptide does not possess inherent uracil-DNA glycosylase activity.

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Section: Methodsmentioning

confidence: 99%

Section: Detection Of Uracil-dna Glycosylase Activity By Using a Uracil-mentioning

confidence: 99%

Escherichia coli nucleoside diphosphate kinase does not act as a uracil-processing DNA repair nuclease

Bennett

Chen²,

Mosbaugh

2004

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

View full text Add to dashboard Cite

show abstract

Purification of Mitochondrial Uracil-DNA Glycosylase Using Ugi-Sepharose Affinity Chromatography

Cited by 1 publication

References 9 publications

Escherichia coli nucleoside diphosphate kinase does not act as a uracil-processing DNA repair nuclease

Escherichia coli nucleoside diphosphate kinase does not act as a uracil-processing DNA repair nuclease

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