Role of the major capsid protein of phage T4 in DNA packaging from structure-function and site-directed mutagenesis studies

Abstract

Heat cleavage of asp-pro peptide bonds was used to probe the primary structures of the Phage T4 major capsid protein precursor, gp23, its mature capsid form gp23 ∗, and a DNA-dependent ATPase, called capsizyme. This analysis suggests that capsizyme is a gp23 ∗∗ resulting from the N-terminal processing found in gp23 ∗ as well as shortening at the C-terminus. Photoaffinity labeling with Azido-ATP and BrU-DNA, followed by heat cleavage, suggests binding sites for these compounds toward the C-terminus of gp23 ∗∗, suggesting localization of functions within the gp23 primary sequence. Site-directed mutagenesis experiments were targeted therefore to the C-terminal end of g23 as well as to its processing sites. N-terminal processing site modification supports the consensus gp21 proteinase cleavage rule, whereas mutagenesis at the C-terminus suggests that the C-terminal alteration is unlikely to result from a gp21-morphogenesis proteinase cleavage. Amino acid replacements in gp23 at newly introduced amber sites reveal a new g23 mutant phenotype, defective partially DNA-filled heads, in support of the hypothesis that gp23 and its products function directly in the DNA packaging mechanism.