Mechanism of Iron-Dependent Repressor (IdeR) Activation and DNA Binding: A Molecular Dynamics and Protein Structure Network Study

Metalloproteins form a major class of enzymes in the living system that are involved in crucial biological functions such as catalysis, redox reactions and as ‘switches’ in signal transductions. Iron dependent repressor (IdeR) is a metal-sensing transcription factor that regulates free iron concentration in Mycobacterium tuberculosis. IdeR is also known to promote bacterial virulence, making it an important target in the field of therapeutics. Mechanistic details of how iron ions modulate IdeR such that it dimerizes and binds to DNA is not understood clearly. In this study, we have performed molecular dynamic simulations and integrated it with protein structure networks to study the influence of iron on IdeR structure and function. A significant structural variation between the metallated and the non-metallated system is observed. Our simulations clearly indicate the importance of iron in stabilizing its monomeric subunit, which in turn promotes dimerization. However, the most striking results are obtained from the simulations of IdeR-DNA complex in the absence of metals, where at the end of 100ns simulations, the protein subunits are seen to rapidly dissociate away from the DNA, thereby forming an excellent resource to investigate the mechanism of DNA binding. We have also investigated the role of iron as an allosteric regulator of IdeR that positively induces IdeR-DNA complex formation. Based on this study, a mechanistic model of IdeR activation and DNA binding has been proposed.

[1]  B. Ames,et al.  The role of heme and iron-sulfur clusters in mitochondrial biogenesis, maintenance, and decay with age. , 2002, Archives of biochemistry and biophysics.

[2]  Illés J. Farkas,et al.  CFinder: locating cliques and overlapping modules in biological networks , 2006, Bioinform..

[3]  M. Bansal,et al.  MolBridge: a program for identifying nonbonded interactions in small molecules and biomolecular structures , 2014 .

[4]  S. Vishveshwara,et al.  Probing the allosteric mechanism in pyrrolysyl-tRNA synthetase using energy-weighted network formalism. , 2011, Biochemistry.

[5]  S. Benkovic,et al.  Flexibility, diversity, and cooperativity: pillars of enzyme catalysis. , 2011, Biochemistry.

[6]  Saraswathi Vishveshwara,et al.  An automated approach to network features of protein structure ensembles , 2013, Protein science : a publication of the Protein Society.

[7]  C. Ratledge.  Iron, mycobacteria and tuberculosis. , 2004, Tuberculosis.

[8]  R. Nussinov,et al.  Allo-network drugs: extension of the allosteric drug concept to protein- protein interaction and signaling networks. , 2013, Current topics in medicinal chemistry.

[9]  M. Perutz.  Stereochemistry of Cooperative Effects in Haemoglobin: Haem–Haem Interaction and the Problem of Allostery , 1970, Nature.

[10]  D. Koshland,et al.  Comparison of experimental binding data and theoretical models in proteins containing subunits. , 1966, Biochemistry.

[11]  V. Hilser,et al.  Rational modulation of conformational fluctuations in adenylate kinase reveals a local unfolding mechanism for allostery and functional adaptation in proteins , 2009, Proceedings of the National Academy of Sciences.

[12]  D. Ringe,et al.  Identification of the primary metal ion-activation sites of the diphtheria tox represser by X-ray crystallography and site-directed mutational analysis , 1996, Nature Structural Biology.

[13]  V. Sobolev,et al.  Flexibility of metal binding sites in proteins on a database scale , 2005, Proteins.

[14]  Gary K. Schoolnik,et al.  ideR, an Essential Gene in Mycobacterium tuberculosis: Role of IdeR in Iron-Dependent Gene Expression, Iron Metabolism, and Oxidative Stress Response , 2002, Infection and Immunity.

[15]  J. Changeux,et al.  ON THE NATURE OF ALLOSTERIC TRANSITIONS: A PLAUSIBLE MODEL. , 1965, Journal of molecular biology.

[16]  A. Amadei,et al.  Thermodynamic and kinetic characterization of a β‐hairpin peptide in solution: An extended phase space sampling by molecular dynamics simulations in explicit water , 2005, Proteins.

[17]  V. Hornak,et al.  Comparison of multiple Amber force fields and development of improved protein backbone parameters , 2006, Proteins.

[18]  H. Berendsen,et al.  Essential dynamics of proteins , 1993, Proteins.

[19]  M. Karplus,et al.  Allostery and cooperativity revisited , 2008, Protein science : a publication of the Protein Society.

[20]  W. Bezwoda,et al.  Haematological and iron-related measurements in active pulmonary tuberculosis. , 2009, Scandinavian journal of haematology.

[21]  T. Darden,et al.  Particle mesh Ewald: An N⋅log(N) method for Ewald sums in large systems , 1993 .

[22]  C. J. Murray,et al.  The adverse effect of iron repletion on the course of certain infections. , 1978, British medical journal.

[23]  H Frauenfelder,et al.  The role of structure, energy landscape, dynamics, and allostery in the enzymatic function of myoglobin , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[24]  Saraswathi Vishveshwara,et al.  Insights into Protein–DNA Interactions through Structure Network Analysis , 2008, PLoS Comput. Biol..

[25]  N. Goodey,et al.  Allosteric regulation and catalysis emerge via a common route. , 2008, Nature chemical biology.

[26]  G. Ciccotti,et al.  Numerical Integration of the Cartesian Equations of Motion of a System with Constraints: Molecular Dynamics of n-Alkanes , 1977 .

[27]  Charles A Laughton,et al.  Essential Dynamics:  A Tool for Efficient Trajectory Compression and Management. , 2006, Journal of chemical theory and computation.

[28]  R. Holmes,et al.  Functional Studies of the Mycobacterium tuberculosis Iron-dependent Regulator* , 2004, Journal of Biological Chemistry.

[29]  Ronald L. Rivest,et al.  Introduction to Algorithms , 1990 .

[30]  S. Vishveshwara,et al.  Identification of side-chain clusters in protein structures by a graph spectral method. , 1999, Journal of molecular biology.

[31]  R. Nussinov,et al.  Is allostery an intrinsic property of all dynamic proteins? , 2004, Proteins.

[32]  Zhou Yu,et al.  The induction of folding cooperativity by ligand binding drives the allosteric response of tetracycline repressor , 2009, Proceedings of the National Academy of Sciences.

[33]  A. Giuliani,et al.  A complexity score derived from principal components analysis of nonlinear order measures , 2001 .

[34]  Saraswathi Vishveshwara,et al.  Rule-based modelling of iron homeostasis in tuberculosis. , 2011, Molecular bioSystems.

[35]  S. Vishveshwara,et al.  Variations in clique and community patterns in protein structures during allosteric communication: investigation of dynamically equilibrated structures of methionyl tRNA synthetase complexes. , 2008, Biochemistry.

[36]  R. Ebright,et al.  Dynamically driven protein allostery , 2006, Nature Structural &Molecular Biology.

[37]  C. Barry,et al.  Getting the iron out , 2005, Nature chemical biology.

[38]  M Paoli,et al.  The stereochemical mechanism of the cooperative effects in hemoglobin revisited. , 1998, Annual review of biophysics and biomolecular structure.

[39]  R. Holmes,et al.  Crystal Structure of the Iron-dependent Regulator from Mycobacterium tuberculosis at 2.0-Å Resolution Reveals the Src Homology Domain 3-like Fold and Metal Binding Function of the Third Domain* , 2000, The Journal of Biological Chemistry.

[40]  Kenneth M Merz,et al.  Structural Survey of Zinc Containing Proteins and the Development of the Zinc AMBER Force Field (ZAFF). , 2010, Journal of chemical theory and computation.

[41]  Shuang-yong Xu,et al.  BstYI bound to noncognate DNA reveals a "hemispecific" complex: implications for DNA scanning. , 2007, Structure.

[42]  S. Vishveshwara,et al.  Allostery and conformational free energy changes in human tryptophanyl‐tRNA synthetase from essential dynamics and structure networks , 2010, Proteins.

[43]  V. Hilser,et al.  The ensemble nature of allostery , 2014, Nature.

[44]  R. Holmes,et al.  Crystal structure of an IdeR-DNA complex reveals a conformational change in activated IdeR for base-specific interactions. , 2004, Journal of molecular biology.