Isolation and partial characterization of murine O6-alkylguanine-DNA-alkyltransferase: comparative sequence and structural properties.

2.50
Hdl Handle:
http://hdl.handle.net/10541/109013
Title:
Isolation and partial characterization of murine O6-alkylguanine-DNA-alkyltransferase: comparative sequence and structural properties.
Authors:
Santibanez-Koref, Mauro F; Elder, Rhoderick H; Fan, Chun-Yang; Cawkwell, Lynn; McKie, J H; Douglas, K T; Margison, Geoffrey P; Rafferty, Joseph A
Abstract:
A cDNA encoding murine O6-alkylguanine-DNA-alkyltransferase (ATase) has been sequenced after isolation from total liver RNA by the polymerase chain reaction using oligonucleotide primers derived from the rat ATase cDNA sequence. Functionally active murine ATase protein has been expressed in Escherichia coli at high levels (about 2% of total protein) and purified to apparent homogeneity (molecular mass 26 kDa). In liquid hybridization experiments, anti-human ATase polyclonal antibodies inhibited human but not rat or mouse ATase, whereas anti-rat polyclonal antibodies inhibited rat and mouse but not human ATase. Both antibodies detected all mammalian ATases tested by western analysis so far. These results indicate some common epitopes and at least one unique human epitope. We compared the amino-acid sequence of the murine ATase with those of other mammalian and bacterial ATases. The proteins of this family all have a large domain (approximately 70 amino acids) of highly conserved residues flanking the sequence PCHRV, which contains the alkyl-accepting cysteine residue of the active site. No evidence was found in the sequences for helix-turn-helix, leucine-zipper, or zinc-finger motifs for DNA recognition and binding. Nuclear localization signals (basic-residue-rich regions) could not be uniquely identified in the mammalian members of the family. Outside of the conserved PCHRV region, there were major differences between prokaryotic and eukaryotic proteins at the primary structure level: there was a series of proline-rich motifs, but these also varied between sequences.
Affiliation:
Department of Cancer Genetics, University of Manchester, United Kingdom.
Citation:
Isolation and partial characterization of murine O6-alkylguanine-DNA-alkyltransferase: comparative sequence and structural properties. 1992, 5 (2):161-9 Mol. Carcinog.
Journal:
Molecular Carcinogenesis
Issue Date:
1992
URI:
http://hdl.handle.net/10541/109013
DOI:
10.1002/mc.2940050212
PubMed ID:
1554415
Type:
Article
Language:
en
ISSN:
0899-1987
Appears in Collections:
All Paterson Institute for Cancer Research

Full metadata record

DC FieldValue Language
dc.contributor.authorSantibanez-Koref, Mauro Fen
dc.contributor.authorElder, Rhoderick Hen
dc.contributor.authorFan, Chun-Yangen
dc.contributor.authorCawkwell, Lynnen
dc.contributor.authorMcKie, J Hen
dc.contributor.authorDouglas, K Ten
dc.contributor.authorMargison, Geoffrey Pen
dc.contributor.authorRafferty, Joseph Aen
dc.date.accessioned2010-08-04T08:58:25Z-
dc.date.available2010-08-04T08:58:25Z-
dc.date.issued1992-
dc.identifier.citationIsolation and partial characterization of murine O6-alkylguanine-DNA-alkyltransferase: comparative sequence and structural properties. 1992, 5 (2):161-9 Mol. Carcinog.en
dc.identifier.issn0899-1987-
dc.identifier.pmid1554415-
dc.identifier.doi10.1002/mc.2940050212-
dc.identifier.urihttp://hdl.handle.net/10541/109013-
dc.description.abstractA cDNA encoding murine O6-alkylguanine-DNA-alkyltransferase (ATase) has been sequenced after isolation from total liver RNA by the polymerase chain reaction using oligonucleotide primers derived from the rat ATase cDNA sequence. Functionally active murine ATase protein has been expressed in Escherichia coli at high levels (about 2% of total protein) and purified to apparent homogeneity (molecular mass 26 kDa). In liquid hybridization experiments, anti-human ATase polyclonal antibodies inhibited human but not rat or mouse ATase, whereas anti-rat polyclonal antibodies inhibited rat and mouse but not human ATase. Both antibodies detected all mammalian ATases tested by western analysis so far. These results indicate some common epitopes and at least one unique human epitope. We compared the amino-acid sequence of the murine ATase with those of other mammalian and bacterial ATases. The proteins of this family all have a large domain (approximately 70 amino acids) of highly conserved residues flanking the sequence PCHRV, which contains the alkyl-accepting cysteine residue of the active site. No evidence was found in the sequences for helix-turn-helix, leucine-zipper, or zinc-finger motifs for DNA recognition and binding. Nuclear localization signals (basic-residue-rich regions) could not be uniquely identified in the mammalian members of the family. Outside of the conserved PCHRV region, there were major differences between prokaryotic and eukaryotic proteins at the primary structure level: there was a series of proline-rich motifs, but these also varied between sequences.en
dc.language.isoenen
dc.subject.meshAmino Acid Sequence-
dc.subject.meshAnimals-
dc.subject.meshBacterial Proteins-
dc.subject.meshBase Sequence-
dc.subject.meshHumans-
dc.subject.meshLiver-
dc.subject.meshMethyltransferases-
dc.subject.meshMice-
dc.subject.meshMolecular Sequence Data-
dc.subject.meshO(6)-Methylguanine-DNA Methyltransferase-
dc.subject.meshProtein Conformation-
dc.subject.meshRats-
dc.subject.meshSequence Alignment-
dc.subject.meshStructure-Activity Relationship-
dc.titleIsolation and partial characterization of murine O6-alkylguanine-DNA-alkyltransferase: comparative sequence and structural properties.en
dc.typeArticleen
dc.contributor.departmentDepartment of Cancer Genetics, University of Manchester, United Kingdom.en
dc.identifier.journalMolecular Carcinogenesisen

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