High affinity of endonuclease VII for the Holliday structure containing one nick ensures productive resolution.
During homologous recombination, genetic information is physically exchanged between parental DNAs via crossing single strands of the same polarity within a four-way DNA junction called a Holliday structure. This process is terminated by the endonucleolytic activity of resolvases, which convert the four-way DNA back to two double strands. To achieve productive resolution, the two subunits of the dimeric enzymes introduce two single-strand cuts positioned symmetrically in opposite strands across the DNA junction. Covalently linked dimers of endonuclease VII from phage T4, whether a homodimer with two or a heterodimer with only one functional catalytic centre, reacted with a synthetic cruciform DNA to form a DNA-enzyme complex immediately after addition of the enzyme. Analysis of the complexes from both reactions revealed that the bound junction contained one nick. While the active homodimer processed this nicked junction consecutively to duplex DNAs by making the second cut, the complex with the heterodimer stayed stable for the whole reaction time. Thus the high affinity of endonuclease VII for the junction containing one nick is part of the mechanism to ensure productive resolution of Holliday structures, by giving the enzyme time to make the second cut, whereupon the complex dissociates into the two duplex DNAs and the free enzyme.
Structural Biology Programme, EMBL, Meyerhofstr. 1, 69117 Heidelberg, Germany.
Rainer P
RP
Birkenbihl
Börries
B
Kemper
D017122:Bacteriophage T4 / Q000201:enzymology*; D001483:Base Sequence; D002384:Catalysis; D004247:DNA / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D024202:Electrophoretic Mobility Shift Assay; D004706:Endodeoxyribonucleases / Q000737:chemistry / Q000378:metabolism*; D004789:Enzyme Activation; D004795:Enzyme Stability; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D021122:Protein Subunits; D011995:Recombination, Genetic; D013379:Substrate Specificity; D013816:Thermodynamics
D021122:Protein Subunits; D004247:DNA; D004706:Endodeoxyribonucleases; C035571:endodeoxyribonuclease VII