The metalloregulatory zinc site in streptococcus pneumoniae AdcR, a Zinc-activated MarR family repressor

Hermes Reyes-Caballero, Alfredo J. Guerra, Faith E. Jacobsen, Krystyna M. Kazmierczak, Darin Cowart, Uma Mahendra Kumar Koppolu, Robert A. Scott, Malcolm E. Winkler, David P. Giedroc

Research output: Contribution to journalArticle

Abstract

Streptococcus pneumoniae D39 AdcR (adhesin competence repressor) is the first metal-sensing member of the MarR (multiple antibiotic resistance repressor) family to be characterized. Expression profiling with a {increment}adcR strain grown in liquid culture (brain-heart infusion) under microaerobic conditions revealed upregulation of 13 genes, including adcR and adcCBA, encoding a high-affinity ABC uptake system for zinc, and genes encoding cell-surface zinc-binding pneumococcal histidine triad (Pht) proteins and AdcAII (Lmb, laminin binding). The {increment}adcR, H108Q and H112Q adcR mutant allelic strains grown in 0.2 mM Zn(II) exhibit a slow-growth phenotype and an approximately twofold increase in cell-associated Zn(II). Apo- and Zn(II)-bound AdcR are homodimers in solution and binding to a 28-mer DNA containing an adc operator is strongly stimulated by Zn(II) with KDNA-Zn=2.4×108M-1 (pH 6.0, 0.2M NaCl, 25°C). AdcR binds two Zn(II) per dimer, with stepwise Zn(II) affinities KZn1 and KZn2 of ≥109M-1 at pH 6.0 and ≥1012M-1 at pH 8.0, and one to three lower affinity Zn(II) depending on the pH. X-ray absorption spectroscopy of the high-affinity site reveals a pentacoordinate N/O complex and no cysteine coordination, the latter finding corroborated by wild type-like functional properties of C30A AdcR. Alanine substitution of conserved residues His42 in the DNA-binding domain, and His108 and His112 in the C-terminal regulatory domain, abolish high-affinity Zn(II) binding and greatly reduce Zn(II)-activated binding to DNA. NMR studies reveal that these mutants adopt the same folded conformation as dimeric wild type apo-AdcR, but fail to conformationally switch upon Zn(II) binding. These studies implicate His42, His108 and H112 as metalloregulatory zinc ligands in S. pneumoniae AdcR.

Original languageEnglish (US)
Pages (from-to)197-216
Number of pages20
JournalJournal of Molecular Biology
Volume403
Issue number2
DOIs
StatePublished - Oct 22 2010
Externally publishedYes

Fingerprint

Microbial Drug Resistance
Streptococcus pneumoniae
Mental Competency
Zinc
DNA
X-Ray Absorption Spectroscopy
Laminin
Alanine
Genes
Cysteine
Up-Regulation
Metals
Ligands
Phenotype
Brain
Growth

Keywords

  • Bacterial pathogen
  • Metalloregulation
  • Zinc homeostasis
  • Zinc sensor
  • Zinc-activated repressor

ASJC Scopus subject areas

  • Molecular Biology
  • Medicine(all)

Cite this

Reyes-Caballero, H., Guerra, A. J., Jacobsen, F. E., Kazmierczak, K. M., Cowart, D., Koppolu, U. M. K., ... Giedroc, D. P. (2010). The metalloregulatory zinc site in streptococcus pneumoniae AdcR, a Zinc-activated MarR family repressor. Journal of Molecular Biology, 403(2), 197-216. https://doi.org/10.1016/j.jmb.2010.08.030

The metalloregulatory zinc site in streptococcus pneumoniae AdcR, a Zinc-activated MarR family repressor. / Reyes-Caballero, Hermes; Guerra, Alfredo J.; Jacobsen, Faith E.; Kazmierczak, Krystyna M.; Cowart, Darin; Koppolu, Uma Mahendra Kumar; Scott, Robert A.; Winkler, Malcolm E.; Giedroc, David P.

In: Journal of Molecular Biology, Vol. 403, No. 2, 22.10.2010, p. 197-216.

Research output: Contribution to journalArticle

Reyes-Caballero, H, Guerra, AJ, Jacobsen, FE, Kazmierczak, KM, Cowart, D, Koppolu, UMK, Scott, RA, Winkler, ME & Giedroc, DP 2010, 'The metalloregulatory zinc site in streptococcus pneumoniae AdcR, a Zinc-activated MarR family repressor', Journal of Molecular Biology, vol. 403, no. 2, pp. 197-216. https://doi.org/10.1016/j.jmb.2010.08.030
Reyes-Caballero, Hermes ; Guerra, Alfredo J. ; Jacobsen, Faith E. ; Kazmierczak, Krystyna M. ; Cowart, Darin ; Koppolu, Uma Mahendra Kumar ; Scott, Robert A. ; Winkler, Malcolm E. ; Giedroc, David P. / The metalloregulatory zinc site in streptococcus pneumoniae AdcR, a Zinc-activated MarR family repressor. In: Journal of Molecular Biology. 2010 ; Vol. 403, No. 2. pp. 197-216.
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