title: Minimum length requirement of the flexible N-terminal translocation subdomain of colicin E3.
abstract: The 315-residue N-terminal T domain of colicin E3 functions in translocation of the colicin across the outer membrane through its interaction with outer membrane proteins including the OmpF porin. The first 83 residues of the T domain are known from structure studies to be disordered. This flexible translocation subdomain contains the TolB box (residues 34 to 46) that must cross the outer membrane in an early translocation event, allowing the colicin to bind to the TolB protein in the periplasm. In the present study, it was found that cytotoxicity of the colicin requires a minimum length of 19 to 23 residues between the C terminus (residue 46) of the TolB box and the end of the flexible subdomain (residue 83). Colicin E3 molecules of sufficient length display normal binding to TolB and occlusion of OmpF channels in vitro. The length of the N-terminal subdomain is critical because it allows the TolB box to cross the outer membrane and interact with TolB. It is proposed that the length constraint is a consequence of ordered structure in the downstream segment of the T domain (residues 84 to 315) that prevents its insertion through the outer membrane via a translocation pore that includes OmpF.
authors.affiliation: Department of Biological Sciences, Purdue University, West Lafayette, IN 47907, USA.
authors.forename: Onkar
authors.initials: O
authors.lastname: Sharma
authors.forename: William A
authors.initials: WA
authors.lastname: Cramer
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000378:metabolism; D001665:Binding Sites / Q000235:genetics; D001693:Biological Transport, Active / Q000502:physiology; D003087:Colicins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000235:genetics / Q000502:physiology; D050296:Microbial Viability; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D033903:Periplasmic Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D018272:Porins / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary
chemical_list: D001425:Bacterial Outer Membrane Proteins; D003087:Colicins; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; C109237:OmpF protein; D033903:Periplasmic Proteins; D018272:Porins; C062094:tolB protein, E coli