1.E.53 The Toxic Hok/Gef Protein (Hok/Gef) Family

Members of the Hok/Gef family include a number of small proteins (about 50 amino acid residues), including the highly similar Gef and RelA proteins of E. coli and several plasmid-encoded proteins such as Hok, FimA, SrnB and PndA (Poulsen et al. 1989). These cell-toxic proteins have N-terminal hydrophobic ''signal'' peptides which traverse the cytoplasmic membrane of the bacteria displaying their C-terminal domains in the periplasm (Poulsen et al. 1991). When overexpressed, these proteins are toxic, killing the bacterial cell from the inside, but their normal physiological functions were not known. They appear to mediate programmed cell death in bacteria, and they function in plasmid maintenance by killing plasmid-free cells via a toxin/unstable (protease-sensitive) antitoxin-dependent mechanism. Members of the Hok/Gef family are found in bacteria and archaea, and the E. coli chromosome encodes at least five hok gene paralogues. Gef forms disulfide-linked homodimers. It has been proposed that they oligomerize to form transmembrane channels or pores that dissipate the membrane potential and thereby kill the cell.  This conclusion has been substantiated (Brielle et al. 2016).

Hok-like proteins are very toxic to most Gram-negative species and also to some extent to Gram-positive bacteria (Gerdes et al. 1997). Induction of Hok leads to loss of the cell membrane potential, arrest of respiration, efflux of small molecules (i.e. Mg²⁺ and ATP), influx of small extracellular molecules (i.e. ONPG) and even influx of periplasmic proteins such as RNase I. By phase contrast microscopy, the cells change morphology to so-called ''ghost-cells'' after induction of Hok protein synthesis. These ghost cells are characterized by condensed cell poles and a centrally located clearing, and resemble the ghost cells (empty cell shells) formed after induction of the lysis gene of φx174. Furthermore, the srnB gene of F complements mutations in the φx174 BS gene , which encodes a holin. Thus the Hok family of proteins bears functional resemblance to holins (TC subclass 1.E). The holins create holes or pores in the inner cell membrane, thereby releasing a phage-encoded endolysin to the periplasm, where the enzyme degrades the cell wall. Thus, the Hok-like proteins kill the cells by mediating irreversible damage to the host cell membrane.