In order to efficiently create new infectious particles, all viruses must solve the problem of specifically identifying their own genomic material from the pool of nucleic acids within the host cell. One mechanism for this specific recognition is the interaction between a packaging signal, a uniquely viral motif within the genomic nucleic acid, and a viral structural protein which then directs the genome for packaging within in the nascent particle. This signal can be a unique sequence of bases that is recognized de novo by the protein, or that can form a secondary structure whose overall shape facilitates the required interaction, or a combination of these two mechanisms.
Large dsDNA viruses create new genomic copies in a continuous strand of dsDNA by a process of rolling circle replication, which need to be cleaved into individual copies and packaged into preformed capsids. These viruses not only need to recognize the packaging signal in the genome, mediated by the small terminase protein, but also coordinate the nuclease activity of the large terminase protein to a specific site to create the nascent end of the genomic DNA that is then loaded, along with the DNA translocation motor, onto the portal pore of the preformed capsid to initiate the packaging process. DNA packaging can take place in several cycles: the initial cycle described above; and subsequent cycles where the packaging motor cleaves the DNA upon packaging of one genomic copy before the motor, bound to the free end of the next genomic copy, is transferred to a new empty capsid and packaging begins anew. Genomic cleavage after completion of each cycle of packaging can be via a headful mechanism where slightly more than one full copy of the genome is packaged, or at specific sequences that generate viruses that contain precisely one copy length of the genome.
SPP1 and SF6 are members of the Siphoviridae for which a wealth of information has been derived from in vitro genetic and biochemical experiments, including the identification of a ~300 bp sequence, the pac site, as necessary for packaging of viral DNA. This information continues to inform a series of structural and biophysical studies of the packaging initiation mechanisms from these, and related thermophilic phages, in the Antson group with aim of understanding the underlying mechanisms for the initiation and loading events in the packaging process. We hope to use this knowledge to design and efficiently construct ‘nanomachines’ for pumping DNA through barriers into natural and synthetic containers such as cells, nano- particles, chambers and channels.