Viral genome packaging terminase cleaves DNA using the canonical RuvC-like two-metal catalysis mechanism

Abstract

Bacteriophages and large dsDNA viruses encode sophisticated machinery to translocate their DNA into a preformed empty capsid. An essential part of this machine, the large terminase protein, processes viral DNA into constituent units utilizing its nuclease activity. Crystal structures of the large terminase nuclease from the thermophilic bacteriophage G20c show that it is most similar to the RuvC family of the RNase H-like endonucleases. Like RuvC proteins, the nuclease requires either Mn2+, Mg2+ or Co2+ ions for activity, but is inactive with Zn2+ and Ca2+. High resolution crystal structures of complexes with different metals reveal that in the absence of DNA, only one catalytic metal ion is accommodated in the active site. Binding of the second metal ion may be facilitated by conformational variability, which enables the two catalytic aspartic acids to be brought closer to each other. Structural comparison indicates that in common with the RuvC family, the location of the two catalytic metals differs from other members of the RNase H family. In contrast to a recently proposed mechanism, the available data do not support binding of the two metals at an ultra-short interatomic distance. Thus we postulate that viral terminases cleave DNA by the canonical RuvC-like mechanism.

Structure of the large terminase from a hyperthermophilic virus reveals a unique mechanism for oligomerization and ATP hydrolysis

“…The nuclease domain resembles the RNase H-like fold ( 36 , 37 ) which, as found for the large terminases of other viruses, differs from other RNase H family proteins by an extended β-sheet and the presence of an auxiliary β-hairpin. Interestingly, the β-hairpin, previously predicted to interact with DNA ( 12 , 38 – 40 ), has an extended hairpin-like loop structure with an α-helix at its tip. The final α-helix (α6) of the nuclease domain is followed by an ordered extended C-terminal loop or ‘arm’ proximal to the ATPase active site that contains a bound sulphate ion, which was present in the crystallization condition.…”

“…The nuclease domain is responsible for cleavage of the genomic DNA concatemer at both the initiation and completion stages of viral DNA packaging ( 11 ). This domain is a member of the RNase H-like endonuclease superfamily and has the highest similarity with the RuvC endonucleases ( 12 – 14 ).…”

Architecture of the herpesvirus genome-packaging complex and implications for DNA translocation

“…with conserved catalytically active residues like D509, E581, D706 and D707 (Selvarajan Sigamani et al, 2013;Xu et al, 2017b). However, the nuclease domains are distal from the central channel (∼45 Å from the exterior of the channel) and the catalytically active residues are oriented towards the sides of the channel in the hexameric ring ( Fig.…”

Porphyrin-Assisted Docking of a Thermophage Portal Protein into Lipid Bilayers: Nanopore Engineering and Characterization

“…51 The X-ray structure of this protein (PDB code 4zjn) revealed a symmetrical arrangement of 12 subunits in a circular assembly. All residues lining the internal channel of this protein have clear electron density, defining the central pore that is only 1.8 nm in diameter at its narrowest portion (Figure 2).…”