Structure of Hjc, a Holliday junction resolvase, from Sulfolobus solfataricus

Abstract

The 2.15-Å structure of Hjc, a Holliday junction-resolving enzyme from the archaeon Sulfolobus solfataricus, reveals extensive structural homology with a superfamily of nucleases that includes type II restriction enzymes. Hjc is a dimer with a large DNA-binding surface consisting of numerous basic residues surrounding the metal-binding residues of the active sites. Residues critical for catalysis, identified on the basis of sequence comparisons and site-directed mutagenesis studies, are clustered to produce two active sites in the dimer, about 29 Å apart, consistent with the requirement for the introduction of paired nicks in opposing strands of the four-way DNA junction substrate. Hjc displays similarity to the restriction endonucleases in the way its specific DNA-cutting pattern is determined but uses a different arrangement of nuclease subunits. Further structural similarity to a broad group of metal͞phosphate-binding proteins, including conservation of active-site location, is observed. A high degree of conservation of surface electrostatic character is observed between Hjc and T4-phage endonuclease VII despite a complete lack of structural homology. A model of the Hjc-Holliday junction complex is proposed, based on the available functional and structural data.

DNA Repair and Genome Maintenance in Bacillus subtilis

“…4). The crystal structure of RecU shows that RecU is similar to type II restriction endonucleases and a Holliday junction endonuclease (Hjc) from the archaeal organism Sulfolobus solfataricus (30,250,289). The overall RecU fold has been described as a "mushroom" with a cap and a stalk (250).…”

The Crystal Structure of D212 from Sulfolobus Spindle-Shaped Virus Ragged Hills Reveals a New Member of the PD-(D/E)XK Nuclease Superfamily

“…Other than this, the increased size of D212 is largely a result of increased loop lengths in regions that connect secondary structural elements and an additional 10 residues at the N terminus. Interestingly, the N-terminal residues are disordered in both D212 (18 disordered residues) and Sso Hjc (8 disordered residues), where these residues have been implicated in DNA binding and are thought to be ordered only upon interaction with DNA (8). Similarly, I-SspI (atypical homing endonuclease) with 151 amino acids is also largely similar, although, consistent with the lower Z score, it exhibits additional differences (Fig.…”

“…However, the characterized archaeal enzymes position their two active site centers on a relatively flat surface (8,34,37) and are predicted to recognize a planar, stacked-X Holliday junction (22,34) like that seen in the structure of T4 endonuclease VII (5). In contrast, the predicted DNA binding channels in D212 run at oblique angles down opposite sides of the dimer.…”

“…Consistent with the lack of functional annotation for D212 and its orthologs, members of the PD-(D/E)XK nuclease superfamily can exhibit extreme variability in their amino acid sequences, making it difficult to recognize highly diverged members (38). Among this family, D212 was found to be most similar to the Holliday junction cleavage enzyme from Sulfolobus solfataricus (Sso Hjc, PDB ID 1HH1), giving a DALI Z-score of 9.0, with an RMSD of 3.0 Å for 115 equivalent residues showing 12% identity (8). Other proteins with significant similarity include additional archaeal Holliday junction cleavage enzymes, an atypical homing endonuclease (I-SspI), and several type II restriction endonucleases.…”

“…This fold and its associated catalytic residues are also found in other classes of nucleases, where they play roles in DNA replication, repair, recombination, and other processes requiring nuclease activity (10). Structural examples include MutH (4); Vsr (56); RecB and -exonuclease (24), which are involved in DNA repair; archaeal Holliday junction cleavage enzymes like Hjc and Hje (8,34,37); TnsA, which is a component of TN7 transposase (19); and the homing endonuclease I-SspI (61). In addition, bioinformatics, biochemical, and/or genetic approaches have identified the PD-(D/E)XK motif in non-long terminal repeat (LTR) retrotransposases (60) and in proteins associated with excisionase activity (10,49,57).…”