title: Zinc coordination is essential for the function and activity of the type II secretion ATPase EpsE.
abstract: The type II secretion system Eps in Vibrio cholerae promotes the extracellular transport of cholera toxin and several hydrolytic enzymes and is a major virulence system in many Gram-negative pathogens which is structurally related to the type IV pilus system. The cytoplasmic ATPase EpsE provides the energy for exoprotein secretion through ATP hydrolysis. EpsE contains a unique metal-binding domain that coordinates zinc through a tetracysteine motif (CXXCX29CXXC), which is also present in type IV pilus assembly but not retraction ATPases. Deletion of the entire domain or substitution of any of the cysteine residues that coordinate zinc completely abrogates secretion in an EpsE-deficient strain and has a dominant negative effect on secretion in the presence of wild-type EpsE. Consistent with the in vivo data, chemical depletion of zinc from purified EpsE hexamers results in loss of in vitro ATPase activity. In contrast, exchanging the residues between the two dicysteines with those from the homologous ATPase XcpR from Pseudomonas aeruginosa does not have a significant impact on EpsE. These results indicate that, although the individual residues in the metal-binding domain are generally interchangeable, zinc coordination is essential for the activity and function of EpsE.
authors.affiliation: Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan.
authors.forename: Chelsea S
authors.initials: CS
authors.lastname: Rule
authors.affiliation: Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan.
authors.forename: Marcella
authors.initials: M
authors.lastname: Patrick
authors.affiliation: Division of Infectious Diseases, Department of Medicine, University of Maryland School of Medicine, Rockville, Maryland.
authors.forename: Jodi L
authors.initials: JL
authors.lastname: Camberg
authors.affiliation: Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan.
authors.forename: Natalie
authors.initials: N
authors.lastname: Maricic
authors.affiliation: Department of Biochemistry, Biomolecular Structure Center, University of Washington, Seattle, Washington.
authors.forename: Wim G
authors.initials: WG
authors.lastname: Hol
authors.affiliation: Department of Microbiology and Immunology, University of Michigan Medical School, Ann Arbor, Michigan. mariasan@umich.edu.
authors.forename: Maria
authors.initials: M
authors.lastname: Sandkvist
mesh_terms: D000251:Adenosine Triphosphatases / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001692:Biological Transport / Q000502:physiology; D002772:Cholera Toxin / Q000378:metabolism*; D036781:Fimbriae Proteins / Q000378:metabolism; D008565:Membrane Proteins / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D011550:Pseudomonas aeruginosa / Q000201:enzymology / Q000378:metabolism; D000069178:Type II Secretion Systems / Q000235:genetics / Q000378:metabolism*; D014734:Vibrio cholerae / Q000201:enzymology / Q000378:metabolism*; D015032:Zinc / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; C079398:EpsE protein, Vibrio cholerae; D008565:Membrane Proteins; D026901:Membrane Transport Proteins; D000069178:Type II Secretion Systems; C082573:xcpR protein, Pseudomonas aeruginosa; D036781:Fimbriae Proteins; D000255:Adenosine Triphosphate; D002772:Cholera Toxin; D000251:Adenosine Triphosphatases; D015032:Zinc
keywords: ATPase; tetracysteine; type II secretion; zinc.