Holliday junction resolving enzymes of archaeal viruses SIRV1 and SIRV2.
In the final stages of genetic recombination, Holliday junction resolving enzymes transform the four-way DNA intermediate into two duplex DNA molecules by introducing pairs of staggered nicks flanking the junction. This fundamental process is apparently common to cells from all three domains of life. Two cellular resolving enzymes from extremely thermophilic representatives of both kingdoms of the domain Archaea, the euryarchaeon Pyrococcus furiosus and the crenarchaeon Sulfolobus solfataricus, have been described recently. Here we report for the first time the isolation, purification and characterization of Holliday junction cleaving enzymes (Hjc) from two archaeal viruses. Both viruses, SIRV1 and SIRV2, infect Sulfolobus islandicus. Their Hjcs both consist of 121 amino acid residues (aa) differing only by 18 aa. Both proteins bind selectively to synthetic Holliday-structure analogues with an apparent dissociation constant of 25 nM. In the presence of Mg(2+) the enzymes produce identical cleavage patterns near the junction. While S. islandicus shows optimal growth at about 80 degrees C, the nucleolytic activities of recombinant SIRV2 Hjc was highest between 45 degrees C and 70 degrees C. Based on their specificity for four-way DNA structures the enzymes may play a general role in genetic recombination, DNA repair and the resolution of replicative intermediates.
Birkenbihl
R P
RP
EMBL, Structural Biology Programme, Meyerhofstr. 1, Heidelberg, 69117, Germany.
Neef
K
K
Prangishvili
D
D
Kemper
B
B
D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D003850:Deoxyribonuclease I / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008274:Magnesium / Q000494:pharmacology; D008969:Molecular Sequence Data; D011485:Protein Binding; D045522:Recombinases; D011995:Recombination, Genetic / Q000235:genetics; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D016944:Sulfolobus / Q000821:virology*; D013696:Temperature; D019895:Transposases / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D014780:Viruses / Q000201:enzymology* / Q000235:genetics
D004268:DNA-Binding Proteins; D045522:Recombinases; D004247:DNA; D019895:Transposases; D003850:Deoxyribonuclease I; D008274:Magnesium