Microbiology Procedure

Index >> Bacteriophages - Part One >> Single Strand Synthesis ( RF to SS )

Single Strand Synthesis ( RF to SS )

	Single Strand Synthesis ( RF to SS ) - The (+) strands released from rolling circles become the genomes of progeny phage particles if protected by the gene V protein. Gene V of Ff phages codes for a binding protein which is analogous to the E. coli unwinding protein or the 32-protein of phage T4.
The protein has a MW of 2,688, and there are about 120,000 copies in the infected cell. In low salt concentration the protein is present in the form of a dimer. As the progeny duplex RF rolls off a single stranded tail, the latter becomes coated with dimers of gene V protein. The DNA protein complexes appear as 1.1 (J.m long and 16 nm wide structures. Each viral DNA molecule binds to about 1,300 molecules of gene V protein. The circular single stranded genome covered with protein now appears as a linear rod.
The protein makes the ssDNA safe from conversion to the RF form. Gene II is essential both for RF replication and SS synthesis. A mutation in the gene prevents both RF -->RF replication and RF--> SS synthesis. The gene II product may be a strand, specific endonuclease required for the initiation of viral strand synthesis. The products of host genes dnaC (D), dnaE (pol III) and polA (poll) are required continuously for phage production. (dnaB and dnaG functions are required only during RF replication). Ligation of ssDNA to give rise to the circular form might be brought about by the host ligase. The origin of viral single strand synthesis is located in the intergenic region.
This region also contains the origin of the complementary strand in SS -->RF formation. The direction of viral strand synthesis is however, in the opposite direction to that of complementary strand synthesis. The growing tail of the rolling circle is cleaved to yield the linear single strand. This endonucleolytic cleavage is presumed to be carried out by the Ff gene U protein.

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