Topologies of complexes containing O6-alkylguanine-DNA alkyltransferase and DNA.
The mutagenic and cytotoxic effects of many alkylating agents are reduced by O(6)-alkylguanine-DNA alkyltransferase (AGT). In humans, this protein not only protects the integrity of the genome, but also contributes to the resistance of tumors to DNA-alkylating chemotherapeutic agents. Here we describe and test models for cooperative multiprotein complexes of AGT with single-stranded and duplex DNAs that are based on in vitro binding data and the crystal structure of a 1:1 AGT-DNA complex. These models predict that cooperative assemblies contain a three-start helical array of proteins with dominant protein-protein interactions between the amino-terminal face of protein n and the carboxy-terminal face of protein n+3, and they predict that binding duplex DNA does not require large changes in B-form DNA geometry. Experimental tests using protein cross-linking analyzed by mass spectrometry, electrophoretic and analytical ultracentrifugation binding assays, and topological analyses with closed circular DNA show that the properties of multiprotein AGT-DNA complexes are consistent with these predictions.
Department of Molecular and Cellular Biochemistry and Center for Structural Biology, University of Kentucky, Lexington, KY 40536, USA.
Claire A
CA
Adams
Manana
M
Melikishvili
David W
DW
Rodgers
Joseph J
JJ
Rasimas
Anthony E
AE
Pegg
Michael G
MG
Fried
D001665:Binding Sites; D002623:Chemistry Techniques, Analytical / Q000379:methods; D004247:DNA / Q000737:chemistry*; D006801:Humans; D046911:Macromolecular Substances / Q000737:chemistry*; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000737:chemistry*; D011487:Protein Conformation
D046911:Macromolecular Substances; D004247:DNA; D019853:O(6)-Methylguanine-DNA Methyltransferase