Why the prince has turned into a frog.
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Biomedical subjects
Publications and source records attributed to C Ganz.
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Defective heads present in extracts of bacteriophage T4 gene 16, 17, or 49 mutant-infected cells have been characterized. All appeared as empty shells when examined by negative-stain electron microscopy and showed essentially the same polypeptide pattern on sodium dodecyl sulfate-acrylamide gels. However, when analyzed by several other methods, gene 16- and 17-defective heads were shown to differ markedly from phage heads present in gene 49-defective extracts. First, the gene 16- and 17-defective structures were found to possess a large number of attached tails (50%, rather than about 5%). Second, they contained less nuclease-resistant deoxyribonucleic acid (DNA) (3 versus 18% of a phage equivalent), had a smaller sedimentation coefficient (240 versus 315S), and a lighter density (1.31 vs. 1.34 g/ml) than gene 49-defective heads. Third, they were not attached to the intracellular DNA pool through a deoxyribonuclease-sensitive linkage. Finally, 8-nm diameter capsomers were clearly revealed on the surface of many gene 16- and 17-defective structures. There was a total of 305 +/- 25 capsomers per particle, which yielded an approximate molecular weight of 84 x 10(6) for these heads. The capsomers were presumably not seen on gene 49-defective heads because of the large amount (18%) of associated DNA. These results support the contention that gene 16- and 17-defective heads, in contrast to gene 49-defective structures, represent abortive rather than intermediate structures in T4 head morphogenesis.
An investigation into the metabolic requirements for maturation of gene 49-defective heads indicated that adenosine triphosphate energy and continued deoxyribonucleic acid (DNA) but not ribonucleic acid synthesis were needed. The fate of DNA present at restrictive temperatures (41.5 C) in tsC9 (gene 49)-infected cells was also examined. After lysis of infected cells, the 12 to 32% deoxyribonuclease-resistant DNA associated with isolated gene 49-defective heads was found to be attached to a deoxyribonuclease-sensitive complex associated with the debris. Pulsechase experiments where (3)H-thymidine was used to label the DNA at 41.5 C suggested that more DNA from this pool was present in phage recovered after rescue of the gene 49 function than could be accounted for by the deoxyribonuclease-resistant portion. Further, when these experiments were repeated with an additional density shift ((15)N(13)C-glucose to (14)N(12)C-glucose), the DNA extracted from phage rescued at 10 min after the temperature shift-down was found to be 90% conserved. These results suggest a model whereby DNA packaging into capsid precursors is separated from DNA replication and the energy from DNA synthesis provides the driving force for packaging. Pulse-chase, temperature-shift experiments with E920g (gene 66) or E920g;tsC9 mutant-infected cells showed that gene (49, 66)-defective heads, which were isolated as small, isometric-shaped unfilled heads, were a precursor to "petite" phage. This suggests that the maturation process is independent of the size and shape of the head membrane. Similar experiments with the double mutant tsC9;amN120 indicate that gene 49-defective heads can also be filled in the absence of tails.
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