Bacteriophage T4D Head Morphogenesis. VIII. DNA-Protein Associations in Intermediate Head Structures That Accumulate in Gene 49 − Mutant-Infected Cells

Abstract

We have utilized the gene 49 − mutant-infected cells of bacteriophage T4D to accumulate large numbers of nucleic acid-protein intermediate head structures. These heads were used as substrates for experiments in the investigations of the mechanism of DNA packaging. Specifically, we have examined: (i) the susceptibility of the DNA in these structures to digestion by a variety of nucleases after a series of increasing temperature pulses from 25 to 100°C, (ii) the physicochemical characteristics of the DNA inside these heads, and (iii) the mechanism by which proteins are displaced from the interior of the head after treatment with basic proteins. We isolated DNA from these gene 49 − heads by use of gradient centrifugation procedures. The DNA had a molecular weight of 8 × 10 6 and a density of 1.697 ± 0.005 g/cm 3 , and it contained a short resistant fraction (SRF) which, when associated with the gene 49 − heads, exhibited AT-protected regions that were not susceptible to micrococcal nuclease digestion. Such a fraction may contain pieces which are important in the initial association of the DNA with the prohead. Exposure of the gene 49 − intermediate capsid structures to basic proteins, such as bovine trypsin inhibitor, lysozyme, and l -polylysine-70, caused a displacement of an amorphous-appearing structure which may be a complex of the gene 49 − DNA and interior components of the capsid (e.g., internal proteins, polyamines). Our general conclusion is that in the gene 49 − intermediate head structures which are only partly filled with DNA, this DNA is held inside the head by strong electrostatic linkages with interior polypeptides and polyamines.