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Selectivity of Metal Ion Binding Specifically Drives the Interaction of a Ni2+-sensor with its operator DNA
No full textTransition metal ions are essential micronutrient for living organisms, being fundamental cofactors of many enzymes that catalyze an enormous variety of biological reactions. At the same time, several metal ions are toxic in their free form for cellular life, demanding a tight regulation of their intracellular trafficking and homeostasis. A paradigmatic example is represented by the human pathogen Helicobacter pylori, which relies on the activity of the nickel-dependent enzymes urease and hydrogenase to survive in the acidic stomach mucosa, where it is responsible of several diseases, such as gastric ulcer and cancer. In this bacterium, homeostasis of essential Ni2+ ions should be tightly controlled to allow growth and colonization. In this processes, NikR, a widespread homo-tetrameric transcription factor controlling the expression of proteins involved in nickel trafficking and/or nickel enzymes, plays a crucial role. The present study investigates the modulation of the interaction between Helicobacter pylori NikR (HpNikR) and DNA, exerted by different concentrations of Ni2+ and/or other metal ions. In particular, the thermodynamic parameters of the interaction between Ni2+, or other divalent metal ions, and HpNikR are determined by using isothermal titration calorimetry (ITC). The metal-dependent capability of HpNikR to bind PureA, the promoter of the urease operon, was determined by using mobility shift assays, DNAse footprinting and ITC. The results provide a congruent description of the factors that determine HpNikR binding to metal ions and DNA and yield clear proofs for activation of HpNikR selectively driven by Ni2+, through a protein conformational rearrangement specifically induced by Ni2+ binding (1-2).
(1) B. Zambelli, M. Bellucci, A. Danielli, V. Scarlato, S. Ciurli; The Ni2+ binding properties of Helicobacter pylori NikR. Chem. Commun., 2007, 35, pp. 3649 - 3651
(2) B. Zambelli, A. Danielli, S. Romagnoli, P. Neyroz, S. Ciurli, V. Scarlato; High-affinity Ni2+ binding selectively promotes binding of Helicobacter pylori NikR to its target urease promoter. J. Mol. Biol., 2008, 383, pp. 1129 - 114
Metal ion selectivity and homeostasis: nickel sensing in the human pathogen Helicobacter pylori
No full textHomeostasis of transition metals is crucial for cellular life and is related to the use of metals as cofactors of many enzymes that catalyze an enormous variety of biological reactions. Living organisms elaborated complex and tightly regulated mechanisms to correctly acquire, utilize and allocate metals, neutralizing their harmful potential. In these processes, a fundamental role is played by metal sensor proteins, which are metal-dependent transcription factors that can select, among several intracellular metal ions, the correct metal cofactor, and mediate the specific cellular response by regulating protein expression. Due to the critical function that metal ion homeostasis generally plays in host-pathogen interactions, metal-sensors are considered fundamental virulence factors, because their deletion diminishes or abrogates the survival and colonization of many pathogens. A paradigmatic example is represented by the human pathogen Helicobacter pylori, which relies on the activity of the nickel-dependent enzymes urease and hydrogenase to survive in the acidic stomach mucosa, where it is responsible of several diseases, such as gastric ulcer and cancer. In this bacterium, nickel homeostasis is governed by NikR, a widespread homo-tetrameric transcription factor controlling the expression of proteins involved in nickel trafficking and/or nickel enzymes, in response to different concentrations of intracellular nickel.
In the present study, the modulation of the interaction between Helicobacter pylori NikR (HpNikR) and DNA, exerted by different concentrations of Ni2+ and/or other metal ions, is investigated. In particular, the thermodynamic parameters of the interaction between Ni2+, or other divalent metal ions, and HpNikR are determined by using isothermal titration calorimetry (ITC). The metal ion-dependent capability of HpNikR to bind PureA, the promoter of the urease operon, was investigated by using mobility shift assays, DNAse footprinting and ITC. The results provide a congruent description of the parameters that determine HpNikR binding to metal ions and DNA and yield clear proofs for activation of HpNikR selectively driven by Ni2+, through a protein conformational rearrangement specifically induced by Ni2+ binding (1-2). A general scheme (see Figure) for the nickel selective HpNikR–DNA interaction is proposed.
(1) B. Zambelli, M. Bellucci, A. Danielli, V. Scarlato, S. Ciurli
The Ni2+ binding properties of Helicobacter pylori NikR
Chem. Commun., 2007, 35, pp. 3649 - 3651
(2) B. Zambelli, A. Danielli, S. Romagnoli, P. Neyroz, S. Ciurli, V. Scarlato
High-affinity Ni2+ binding selectively promotes binding of Helicobacter pylori NikR to its target urease promoter
J. Mol. Biol., 2008, 383, pp. 1129 - 114
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