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Activation of latent proteinases ensures that the timing of proteolysis is regulated precisely, a process that generally involves proteolytic excision of a pro-region or a tightly bound inhibitor. Here we define the activation mechanism for Lit, a dormant suicide proteinase in Escherichia coli K-12. Previous work has shown that Gol, a short sequence within the major capsid protein gp23, activates Lit during the latter stages of T4 phage infection. This results in cell death and exclusion of the phage from the culture. The Lit site specifically cleaves the host translation factor EF-Tu (elongation factor Tu) after it has formed a weak complex with Gol, which can be supplied as a 29-residue peptide. Gol is absolutely required for Lit activation. but its role in proteolysis is unknown. Using a purified three-component system and kinetic analysis, we demonstrate that under physiological conditions Lit hydrolyzes its substrate very slowly (k(cat) of approximately 1 s(-1)). Given the abundance of EF-Tu in the cell, this finding is consistent with a cell-killing mechanism in which a few cleaved EF-Tu proteins are able block translating ribosomes from functioning. We also demonstrate that less than half of the 29 Gol residues are needed for Lit activation and that the role of the peptide is not to provide catalytic groups but to influence catalysis indirectly through stabilization of the ternary Lit.Gol.EF-Tu complex. Hence, phage-elicited suicide of E. coli K-12 by Lit is a variant form of "cofactor-induced activation," a mechanism of protease activation that has only been documented previously in pathogen subversion of mammalian hemostasis cascades.

Original publication




Journal article


J Biol Chem

Publication Date





112 - 117


Amino Acid Sequence, Bacteriophage T4, Endopeptidases, Escherichia coli, Escherichia coli Proteins, Gene Expression Regulation, Bacterial, Hydrolysis, Kinetics, Membrane Proteins, Molecular Sequence Data, Mutagenesis, Peptide Elongation Factor Tu, Substrate Specificity