The linear double-stranded DNA molecules in virions are generated by nicking of concatemeric intracellular DNA by terminase, the DNA packaging enzyme. Staggered nicks are introduced at cosN to generate the cohesive ends of virion DNA. After nicking, the cohesive ends are separated by terminase; terminase bound to the left end of the DNA to be packaged then binds the empty protein shell, i.e., the prohead, and translocation of DNA into the prohead occurs. cosB, a site adjacent to cosN, is a terminase binding site. cosB facilitates the rate and fidelity of the cosN cleavage reaction by serving as an anchoring point for gpNu1, the small subunit of terminase. cosB is also crucial for the formation of a stable terminase-DNA complex, called complex I, formed after cosN cleavage. The role of complex I is to bind the prohead. Mutations in cosB affect both cosB functions, causing mild defects in cosN cleavage and severe packaging defects. R2؊ R1 ؊ DNA. Observations on the adenosine triphosphatase activities and protease susceptibilities of gpNu1 of the Nu1ms1 and Nu1ms3 terminases indicate that the conformation of gpNu1 is altered in the suppressing terminases.Phage is one of many DNA viruses that produce concatemeric DNA during lytic development, leading to the requirement that unit-length, virion chromosomes be generated by cleavage of the concatemer. All of the viruses so far studied that process concatemeric DNA also construct a preformed capsid shell, the procapsid or prohead, into which the DNA is translocated; cutting the concatemer accompanies the packaging process. virion DNA is a linear duplex 48,502 bp in length, with 12-base, complementary cohesive ends at the 5Ј ends of the strands. The cohesive ends allow cyclization of the chromosome upon injection into a cell. Regeneration of the cohesive ends occurs through the introduction, by the phageencoded enzyme terminase, of nicks staggered by 12 bp at cosN, the site containing the cohesive end sequence. cosN is just one of three DNA signals in a larger segment called cos. cos also contains cosB, a terminase binding site required for initiation of packaging, and cosQ, a site required for termination (Fig. 1).Terminase consists of a small subunit, gpNu1, the 21-kDa product of the Nu1 gene, and a large subunit, gpA, the 74-kDa product of the A gene (Fig. 1). Terminase is a bivalent DNA binding protein. The endonuclease activity of terminase, which nicks cosN, is in gpA (15,32,33), and the cosB binding activity is in gpNu1 (36). Mutations in gpA that specifically inactivate the endonuclease activity have been identified in the C-terminal third of gpA (15,26,27). Binding of cosB presumably occurs through a putative helix-turn-helix DNA binding domain at the amino terminus of gpNu1 (5).Because cosN shows partial rotational symmetry, cosN is presumably recognized and nicked by symmetrically disposed gpAs (5, 7). There are three gpNu1 binding sites in cosB: R3, R2, and R1 (36). Between R3 and R2 is I1, a binding site for the Escherichia coli DNA bending protein IHF (28,...