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

Rui-Gang Xu,Alfred A Antson,Huw T Jenkins,Evgeny Klimuk,Maria Chechik,Elena V Blagova,Leonid Minakhin,Konstantin Severinov,Anna Lopatina,Sandra J Greive

doi:10.1093/nar/gkw1354

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.

Highlights

The large terminase protein is a key component of the DNA packaging machinery in tailed bacteriophages and evolutionarily related herpes viruses [1,2]
As in other members of the RNase H-like endonucleases, a cluster of carboxylic acids is contributed to the active site by strands ␤3, ␤4 and ␤6, helix ␣5 and loops L0–L3
These residues were shown to be critical for bacteriophage function, DNA packaging or nuclease activities in bacteriophages T4 [17,38] and SPP1 [16,39]

Summary

Introduction

The large terminase protein is a key component of the DNA packaging machinery in tailed bacteriophages and evolutionarily related herpes viruses [1,2]. Unlike phage ␭ where the nuclease cuts at the specific cosN site [8], in other phages such as T4, SPP1 and P22, only the first cut is made at a specific sequence close to the packaging (pac) site while the second cut is nonsequence specific [9,10,11]. This second, or headful, cleavage event is made after around 102 to 110% of a genome length DNA has been packaged into the procapsid [12]

Methods

Results

Conclusion