Model for DNA packaging into bacteriophage T4 heads

L W Black, D J Silverman
1978 Journal of Virology  
The mechanism of DNA packaging into bacteriophage T4 heads in vivo was investigated by glucosylation of hydroxymethlycytosine residues in a conditionally glucose-deficient host. Cytoplasmic DNA associated with partially packaged ts49 heads can be fully glucosylated, whereas DNA already packaged into these heads is shown to be resistant to glucosylation. After temperature shift and completion of arrested packaging into the reversible temperature-sensitive ts49 head, the structure of the DNA in
more » ... ure of the DNA in the mature ts49 phage was investigated by restriction enzyme digestion, autoradiography, and other techniques. Such mature DNA appears to be fully glucosylated along part of its length and nonglucosylated on the remainder. Its structure suggests that the DNA is run into the head linearly and unidirectionally from one mature end and that there is little sequence specificity in that portion of the T4 DNA which first enters the capsid. This technique should be useful in investigation of the three-dimensional structure of first-and last-packaged DNA within the head; preliminary studies including autoradiography of osmotically shocked phage suggest that the DNA which first enters the head is deposited toward the center of the capsid and that the end of the DNA which first enters the head exits first upon injection. In conjunction with studies of the structure of condensed DNA, the positions and functions of T4 capsid proteins in DNA packaging, and the order of T4 packaging functions [], the features described above suggest the following model: the first DNA end is fixed to the proximal apex of the head at p20 and the DNA is then pumped into the head enzymatically by proteins (p20 + p17) which induce torsion in the DNA molecule. The structure of condensed DNA within viruses and the mechanism of its condensation and decondensation are of interest for understanding higher-order DNA structural organization and intracellular mobilization. A plausible model for the general organization of DNA within bacteriophage heads has come from electron micrographs of highly condensed DNA suddenly released from capsids (31) and X-ray analysis of isometric phage heads in solution (8). These investigations have suggested that DNA within the phage head is tightly wound as a coaxial spool (see Fig. 10i , which was taken from reference 8). How DNA is tightly packaged into the phage head from the metabolically active and extended DNA within the cell and how it can subsequently be rapidly delivered (within seconds in the case of phage T4) from the head upon injection into the infected cell are not understood. It had been proposed that the DNA is packaged into the head by a pulling mechanism arising from cleavage or exit of core proteins from the prehead (4, 20), by expansion of the prehead (13, 32), or in a spontaneous process involving charge neutralization by small molecules and/or attachment to capsid inner-surface proteins (12). Any proposed mechanism should attempt to account for the energetics and the reversibility of DNA condensation. In this paper a model is advanced for DNA condensation which appears to be consistent with structural analyses of condensed DNA (8, 31) , with experiments reported here on the dynamics of DNA packaging into the phage T4 head and the structure of this DNA, with structural protein locations in the bacteriophage T4 capsid and their functions in DNA packaging (14, 28) , and with T4 functions known to be required for packaging and their order of function (14; C. L. Hsiao and L. W. Black, Virology, in press). MATERIALS AND METHODS Bacteria and bacteriophage. Escherichia coli DF2000 (phosphoglucose isomerase-, glucose-6-P-de-643 on May 7, 2020 by guest
doi:10.1128/jvi.28.2.643-655.1978 fatcat:jhzkizlilbfadaebeqnjrhhrg4