Crystal structure of the DNA-recognition component of the bacterial virus Sf6 genome-packaging machine

Abstract

In herpesviruses and many bacterial viruses, genome-packaging is a precisely mediated process fulfilled by a virally encoded molecular machine called terminase that consists of two protein components: A DNA-recognition component that defines the specificity for packaged DNA, and a catalytic component that provides energy for the packaging reaction by hydrolyzing ATP. The terminase docks onto the portal protein complex embedded in a single vertex of a preformed viral protein shell called procapsid, and pumps the viral DNA into the procapsid through a conduit formed by the portal. Here we report the 1.65 Å resolution structure of the DNA-recognition component gp1 of the Shigella bacteriophage Sf6 genomepackaging machine. The structure reveals a ring-like octamer formed by interweaved protein monomers with a highly extended fold, embracing a tunnel through which DNA may be translocated. The N-terminal DNA-binding domains form the peripheral appendages surrounding the octamer. The central domain contributes to oligomerization through interactions of bundled helices. The C-terminal domain forms a barrel with parallel beta-strands. The structure reveals a common scheme for oligomerization of terminase DNA-recognition components, and provides insights into the role of gp1 in formation of the packaging-competent terminase complex and assembly of the terminase with the portal, in which ring-like protein oligomers stack together to form a continuous channel for viral DNA translocation.terminase | DNA-packaging | oligomer | bacteriophage | protein:DNA interaction I n many tailed double-stranded DNA (dsDNA) bacteriophages and herpesvirus, the newly synthesized viral DNA in the infected host cell is present as a concatemer that is comprised of multiple copies of unit-length genome. The concatemeric DNA is packaged into a preformed capsid precursor termed procapsid through a channel formed by a portal protein complex embedded in a unique vertex of the procapsid (1-5). The DNApackaging process involves a precisely coordinated sequence of molecular events, driven by a molecular machine called terminase consisting of two proteins: A DNA-recognition component and a catalytic component, also known as the "small" and "large" subunit, respectively. The terminase specifically binds to concatemeric viral DNA through the DNA-recognition component and cleaves it using the catalytic component, generating a new terminus for the DNA. This terminus is threaded through the portal protein channel embedded in the procapsid, and the catalytic component pumps the DNA into the procapsid in an ATP-dependent manner. When an appropriate amount of DNA is inserted, the terminase cuts the DNA again, dissociates from the procapsid, and can start the next cycle of DNA-packaging. Many of these features in DNA-packaging are also shared by adenoviruses (6). The x-ray structures of the catalytic component from bacteriophage T4 and the portals from various phages were previously described (7-10). However, it is not known how the terminases are assembled and h...

The 1.58 Å resolution structure of the DNA-binding domain of bacteriophage SF6 small terminase provides new hints on DNA binding

“…The fourth helix acts as a linker between the DNA-binding domain and the oligomerization domain (Bü ttner et al, 2012). The same feature is found in the DBD of Shigella bacteriophage Sf6 small terminase, in which the fourth helix following the HTH links the DBD to the oligomerization domain (Zhao et al, 2010). Likewise, in bacteriophage the fourth helix following the DBD domain ( D) has also been proposed to link the DBD to the oligomerization domain (de Beer et al, 2002;Bain et al, 2001).…”

“…To date the crystal structures of a few terminases have been determined, but none of them are in a complex with DNA ( de Beer et al, 2002;Zhao et al, 2010;Roy & Cingolani, 2012). The recent determination of the 4 Å resolution crystal structure of full-length SF6 G1P has allowed the proposal of a DNA-binding model characterized by the DNA wrapping around the outside of a nine-subunit G1P oligomer as in the nucleosome (Bü ttner et al, 2012).…”

Structural basis for DNA recognition and loading into a viral packaging motor

“…Three-dimensional information on small terminases is limited to the cryo-EM structure of phage P22 small terminase (12), the NMR structure of the DNA-binding domain of phage λ gpNu1 (13), and the crystal structure of phage Sf6 small terminase (14). In the absence of accurate three-dimensional data for all three motor components of one particular phage, mapping of functional information to the structure and modeling molecular interactions between individual components is challenging.…”

Compression of the DNA substrate by a viral packaging motor is supported by removal of intercalating dye during translocation

“…High-resolution structures of all of the conserved motor components found among tailed dsDNA bacteriophages have been determined (2)(3)(4)(5). The small bacteriophage T4 terminase protein gp16 is required for cutting and packaging the replicative DNA concatemer in vivo but is nonessential and inhibitory for linear DNA packaging in vitro (6).…”