The DNA-Delay Mutants of Bacteriophage T4

Abstract

observations, taken together with those of others, suggest that these products are necessary for normal DNA synthesis and that they interact with each other and a bacterial component probably at the membrane. MATERIALS AND METHODS Phage and bacterial strains. All T4 phage mutants used in this study came from the California Institute of Technology collection under the care of W. Wood. The designations of these mutants are indicated in the gene maps shown in Fig. 1. Three E. coli strains lacking an am suppressor S/6, 594, and B/5, as well as two strains containing an am suppressor, CR63 and 011', were used. E. coli 594 (4) and CR63 (1) were derived from E. coli K12, whereas S/6, B/5, and 011' were derived from E. coli B (11, 31). Media. H-broth was used for growth and suspension of phage and bacteria. EHA bottom and top agar were used for plaque assays or for plating bacteria (25). M9 medium (6) was used for growth of infected cells in DNA-labeling experiments. Preparation of phage stocks. The phage stocks used were clonally derived from single plaques. High titer phage stocks were prepared by a plate lysate method adapted from that described by Edgar (9). Phage lysates used for [-'H]thymidine incorporation experiments were prepared on M9 plates or in M9 liquid medium without thymidine. Preparation of bacterial plating cultures. Fresh log phase cultures of bacteria were used for phage assays. These were prepared by diluting a fresh overnight culture 1:1,000 in H-broth and growing the 860

Mutational analysis of a type II topoisomerase cleavage site: distinct requirements for enzyme and inhibitors.

“…The physiologic roles of type II topoisomerases include introduction and removal of DNA supercoils (Liu and Wang, 1987;Drlica, 1990;Reece and Maxwell, 1991), segregation of DNA molecules during mitosis (Uemura et al, 1987;Holm et al, 1989;Downes et al, 1991) and possibly attachment of DNA loops to eukaryotic chromosomal Oxford University Press scaffolds (Berrios et al, 1985;. The bacteriophage T4 type II topoisomerase, which is the focus of this study, is required for normal phage DNA replication and transcription (Yegian et al, 1971;Mufti and Bernstein, 1974;McCarthy et al, 1976;Mosig et al, 1983) and may also be important for membrane -DNA association and for packaging DNA from the complex networks that are generated during the course of the infective cycle.…”

A nascent peptide is required for ribosomal bypass of the coding gap in bacteriophage T4 gene 60

“…Gene 60 encodes the 18 kd subunit required for tight complex formation of the three subunits and allows reconstitution of the ATP-dependent catalytic topoisomerase activity. T4 amber mutants defective for gene 60 display a cold-sensitive phenotype as well as a delay in the onset of phage DNA replication (Mufti and Bernstein, 1974).…”

The genome of bacteriophage T4.

“…3). Defects in genes 39, 52, 58 to 61, and 60 delay the onset of DNA synthesis at 370C, but prevent DNA synthesis entirely at 250C (141). The functions of these genes remain intriguingly obscure.…”