Zhen Ma

Publications (4)25 Total impact

• Article: Origins of specificity and cross-talk in metal ion sensing by Bacillus subtilis Fur.

  Zhen Ma, Melinda J Faulkner, John D Helmann
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  ABSTRACT: Fur (ferric uptake regulator) is the master regulator of iron homeostasis in many bacteria, but how it responds specifically to Fe(II) in vivo is not clear. Biochemical analyses of Bacillus subtilis Fur (BsFur) reveal that in addition to Fe(II), both Zn(II) and Mn(II) allosterically activate BsFur-DNA binding. Dimeric BsFur co-purifies with site 1 structural Zn(II) (Fur(2) Zn(2) ) and can bind four additional Zn(II) or Mn(II) ions per dimer. Metal ion binding at previously described site 3 occurs with highest affinity, but the Fur(2) Zn(2) :Me(2) form has only a modest increase in DNA binding affinity (approximately sevenfold). Metallation of site 2 (Fur(2) Zn(2) :Me(4) ) leads to a ∼ 150-fold further enhancement in DNA binding affinity. Fe(II) binding studies indicate that BsFur buffers the intracellular Fe(II) concentration at ∼ 1 μM. Both Mn(II) and Zn(II) are normally buffered at levels insufficient for metallation of BsFur site 2, thereby accounting for the lack of cross-talk observed in vivo. However, in a perR mutant, where the BsFur concentration is elevated, BsFur may now use Mn(II) as a co-repressor and inappropriately repress iron uptake. Since PerR repression of fur is enhanced by Mn(II), and antagonized by Fe(II), PerR may co-regulate Fe(II) homeostasis by modulating BsFur levels in response to the Mn(II)/Fe(II) ratio.
  Molecular Microbiology 10/2012; · 5.01 Impact Factor
• Article: Derepression of the Bacillus subtilis PerR peroxide stress response leads to iron deficiency.

  Melinda J Faulkner, Zhen Ma, Mayuree Fuangthong, John D Helmann
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  ABSTRACT: The Bacillus subtilis PerR repressor regulates the adaptive response to peroxide stress. The PerR regulon includes the major vegetative catalase (katA), an iron storage protein (mrgA), an alkylhydroperoxide reductase (ahpCF), a zinc uptake system (zosA), heme biosynthesis enzymes (hemAXCDBL), the iron uptake repressor (fur), and perR itself. A perR null strain is resistant to hydrogen peroxide, accumulates a porphyrin-like compound, and grows very slowly. The poor growth of the perR mutant can be largely accounted for by the elevated expression of two proteins: the KatA catalase and Fur. Genetic studies support a model in which poor growth of the perR null mutant is due to elevated repression of iron uptake by Fur, exacerbated by heme sequestration by the abundant catalase protein. Analysis of the altered-function allele perR991 further supports a link between PerR and iron homeostasis. Strains containing perR991 are peroxide resistant but grow nearly as well as the wild type. Unlike a perR null allele, the perR991 allele (F51S) derepresses KatA, but not Fur, which likely accounts for its comparatively rapid growth.
  Journal of bacteriology 12/2011; 194(5):1226-35. · 3.94 Impact Factor
• Source

  Available from: John D Helmann

  Article: Sequential binding and sensing of Zn(II) by Bacillus subtilis Zur.

  Zhen Ma, Scott E Gabriel, John D Helmann
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  ABSTRACT: Bacillus subtilis Zur (BsZur) represses high-affinity zinc-uptake systems and alternative ribosomal proteins in response to zinc replete conditions. Sequence alignments and structural studies of related Fur family proteins suggest that BsZur may contain three zinc-binding sites (sites 1-3). Mutational analyses confirm the essential structural role of site 1, while mutants affected in sites 2 and 3 retain partial repressor function. Purified BsZur binds a maximum of two Zn(II) per monomer at site 1 and site 2. Site 3 residues are important for dimerization, but do not directly bind Zn(II). Analyses of metal-binding affinities reveals negative cooperativity between the two site 2 binding events in each dimer. DNA-binding studies indicate that BsZur is sequentially activated from an inactive dimer (Zur(2):Zn(2)) to a partially active asymmetric dimer (Zur(2):Zn(3)), and finally to the fully zinc-loaded active form (Zur(2):Zn(4)). BsZur with a C84S mutation in site 2 forms a Zur(2):Zn(3) form with normal metal- and DNA-binding affinities but is impaired in formation of the Zur(2):Zn(4) high affinity DNA-binding state. This mutant retains partial repressor activity in vivo, thereby supporting a model in which stepwise activation by zinc serves to broaden the physiological response to a wider range of metal concentrations.
  Nucleic Acids Research 08/2011; 39(21):9130-8. · 8.03 Impact Factor
• Source

  Available from: John D Helmann

  Article: Identification of altered function alleles that affect Bacillus subtilis PerR metal ion selectivity.

  Zhen Ma, Jin-Won Lee, John D Helmann
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  ABSTRACT: Bacillus subtilis PerR is a Fur family repressor that senses hydrogen peroxide by metal-catalyzed oxidation. PerR contains a structural Zn(II) ion (Site 1) and a regulatory metal binding site (Site 2) that, upon association with either Mn(II) or Fe(II), allosterically activates DNA binding. In addition, a third less conserved metal binding site (Site 3) is present near the dimer interface in several crystal structures of homologous Fur family proteins. Here, we show that PerR proteins with substitutions of putative Site 3 residues (Y92A, E114A and H128A) are functional as repressors, but are unexpectedly compromised in their ability to sense H(2)O(2). Consistently, these mutants utilize Mn(II) but not Fe(II) as a co-repressor in vivo. Metal titrations failed to identify a third binding site in PerR, and inspection of the PerR structure suggests that these residues instead constitute a hydrogen binding network that modulates the architecture, and consequently the metal selectivity, of Site 2. PerR H128A binds DNA with high affinity, but has a significantly reduced affinity for Fe(II), and to a lesser extent for Mn(II). The ability of PerR H128A to bind Fe(II) in vivo and to thereby respond efficiently to H(2)O(2) was restored in a fur mutant strain with elevated cytosolic iron concentration.
  Nucleic Acids Research 03/2011; 39(12):5036-44. · 8.03 Impact Factor