Identification and characterization of a high-affinity zinc uptake system in Neisseria gonorrhoeae.
A search of the gonococcal genome database using the known zinc-binding protein (znuA) sequences from Escherichia coli and Haemophilus influenzae identified an open reading frame encoding a putative gonococcal ZnuA. The consensus amino acid sequence of this open reading frame possessed a characteristic 30-amino acid histidine-rich metal-binding motif (repetitive HDH sequence) containing 43% histidine and 37% aspartic acid and glutamic acid. Subsequently, two adjacent open reading frames with homology to E. coli and H. influenzae znuB and znuC were located upstream of znuA. When partially purified from sonicated cell-free supernatants by CM-Sepharose chromatography, the mature gonococcal ZnuA had an estimated molecular mass of 38 kDa by SDS-PAGE. The presence of a DNA sequence encoding a 19-amino acid signal peptide and the solubility of the mature ZnuA suggested that this protein was located in the periplasm. Inactivation of the Neisseria gonorrhoeae F62 znuA by insertional mutagenesis resulted in a mutant that had a growth rate lower than that of the wild-type parent strain and that required high concentrations of ZnCl2 (> or = 200 microM) for optimal growth. Using a chemically defined agar medium, the gonococcal ZnuA mutant grew only in the presence of Zn(2+), whereas Mg(2+), Ca(2+), Ni(2+), Fe(2+), Cu(2+), Mn(2+) and Cd(2+) had either no effect or were growth inhibitory.
Chen
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Division of AIDS, STD, and TB Laboratory Research, National Center for Infectious Diseases, Centers for Disease Control and Prevention, Mail stop: G-39, 1600 Clifton Rd., Atlanta, GA 30333, USA. cycl@cdc.gov
Morse
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D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D001692:Biological Transport; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*; D027682:Cation Transport Proteins; D003001:Cloning, Molecular; D030561:Databases, Nucleic Acid; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D009344:Neisseria gonorrhoeae / Q000187:drug effects / Q000235:genetics* / Q000254:growth & development / Q000378:metabolism*; D016366:Open Reading Frames / Q000235:genetics; D017385:Sequence Homology; D015032:Zinc / Q000378:metabolism* / Q000494:pharmacology
D001426:Bacterial Proteins; D002352:Carrier Proteins; D027682:Cation Transport Proteins; C450044:ZnuA protein, Neisseria gonorrhoeae; D015032:Zinc