Terminal Proteins Enable Initiation at the Ends of Viral DNAs

KEY CONCEPT
- A terminal protein binds to the 5′ end of DNA and provides a cytidine nucleotide with a 3′–OH end that primes replication.
An example of initiation at a linear end is provided by adenovirus and Ф29 DNAs, which actually replicate from both ends using the mechanism of strand displacement illustrated in FIGURE 1. The same events can occur independently at either end. Synthesis of a new strand starts at one end, displacing the homologous strand that was previously paired in the duplex. When the replication fork reaches the other end of the molecule, the displaced strand is released as a free single strand. It is then replicated independently; this requires the formation of a duplex origin by base pairing between some short complementary sequences at the ends of the molecule.

FIGURE 1. Adenovirus DNA replication is initiated separately at the two ends of the molecule and proceeds by strand displacement.
In several viruses that use such mechanisms, a protein is found covalently attached to each 5′ end. In the case of adenovirus, a terminal protein is linked to the mature viral DNA via a phosphodiester bond to serine, as indicated in FIGURE 2..

FIGURE 2. The 5′ terminal phosphate at each end of adenovirus DNA is covalently linked to serine in the 55-kD Ad-binding protein.
How does the attachment of the protein overcome the initiation problem? The terminal protein has a dual role: It carries a cytidine nucleotide that provides the primer –OH, and it is associated with DNA polymerase. In fact, linkage of terminal protein to a nucleotide is undertaken by DNA polymerase in the presence of adenovirus DNA. This suggests the model illustrated in FIGURE 3. The complex of polymerase and terminal protein, bearing the priming C nucleotide, binds to the end of the adenovirus DNA. The free 3′–OH end of the C nucleotide is used to prime the elongation reaction by the DNA polymerase. This generates a new strand whose 5′ end is covalently linked to the initiating C nucleotide. (The reaction actually involves displacement of protein from DNA rather than binding de novo. The 5′ end of adenovirus DNA is bound to the terminal protein that was used in the previous replication cycle. The old terminal protein is displaced by the new terminal protein for each new replication cycle.)

FIGURE 3. Adenovirus terminal protein binds to the 5′ end of DNA and provides a C–OH end to prime synthesis of a new DNA strand.
Terminal protein binds to the region located between 9 and 18 bp from the end of the DNA. The adjacent region, between positions 17 and 48, is essential for the binding of a host protein, nuclear factor I, which is also required for the initiation reaction. The initiation complex may therefore form between positions 9 and 48, a fixed distance from the end of the DNA.