Molecular modeling and functional analysis of the AtoS-AtoC two-component signal transduction system of Escherichia coli.

The AtoS-AtoC two-component signal transduction system positively regulates the expression of the atoDAEB operon in Escherichia coli. Upon acetoacetate induction, AtoS sensor kinase autophosphorylates and subsequently phosphorylates, thereby activating, the response regulator AtoC. In a previous work we have shown that AtoC is phosphorylated at both aspartate 55 and histidine73. In this study, based on known three-dimensional structures of other two component regulatory systems, we modeled the 3D-structure of the receiver domain of AtoC in complex with the putative dimerization/autophosphorylation domain of the AtoS sensor kinase. The produced structural model indicated that aspartate 55, but not histidine 73, of AtoC is in close proximity to the conserved, putative phosphate-donor, histidine (H398) of AtoS suggesting that aspartate 55 may be directly involved in the AtoS-AtoC phosphate transfer. Subsequent biochemical studies with purified recombinant proteins showed that AtoC mutants with alterations of aspartate 55, but not histidine 73, were unable to participate in the AtoS-AtoC phosphate transfer in support of the modeling prediction. In addition, these AtoC mutants displayed reduced DNA-dependent ATPase activity, although their ability to bind their target DNA sequences in a sequence-specific manner was found to be unaltered.

[1]  W. D. Nunn,et al.  Regulation of the ato operon by the atoC gene in Escherichia coli , 1987, Journal of bacteriology.

[2]  T. Mizuno,et al.  Phosphotransfer circuitry of the putative multi‐signal transducer, ArcB, of Escherichia coli: in vitro studies with mutants , 1995, Molecular microbiology.

[3]  U. K. Laemmli,et al.  Cleavage of Structural Proteins during the Assembly of the Head of Bacteriophage T4 , 1970, Nature.

[4]  B Magasanik,et al.  Phosphorylation of nitrogen regulator I of Escherichia coli induces strong cooperative binding to DNA essential for activation of transcription. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[5]  D. Higgins,et al.  T-Coffee: A novel method for fast and accurate multiple sequence alignment. , 2000, Journal of molecular biology.

[6]  D. Kyriakidis,et al.  Modulation of ornithine decarboxylase activity in Escherichia coli by positive and negative effectors. , 1978, Proceedings of the National Academy of Sciences of the United States of America.

[7]  D. Kyriakidis,et al.  Involvement of the AtoS-AtoC signal transduction system in poly-(R)-3-hydroxybutyrate biosynthesis in Escherichia coli. , 2006, Biochimica et biophysica acta.

[8]  H. Towbin,et al.  Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. , 1979, Proceedings of the National Academy of Sciences of the United States of America.

[9]  Eleni P. Mimitou,et al.  Phosphorylation activity of the response regulator of the two-component signal transduction system AtoS-AtoC in E. coli. , 2005, Biochimica et biophysica acta.

[10]  Qingping Xu,et al.  The yeast YPD1/SLN1 complex: insights into molecular recognition in two-component signaling systems. , 2003, Structure.

[11]  D. Bruns,et al.  A sensitive and precise isotopic assay of ATPase activity. , 1978, Analytical biochemistry.

[12]  T. Mizuno,et al.  Transmembrane signal transduction and osmoregulation in Escherichia coli: II. The osmotic sensor, EnvZ, located in the isolated cytoplasmic membrane displays its phosphorylation and dephosphorylation abilities as to the activator protein, OmpR. , 1990, Journal of biochemistry.

[13]  A. F. Whiting.  Identification , 1960, Australian Water Bugs. (Hemiptera - Heteroptera, Gerromorpha & Nepomorpha).

[14]  R. Dixon,et al.  Domain Architectures of σ54-Dependent Transcriptional Activators , 2003 .

[15]  Seok-Yong Lee,et al.  Regulation of the transcriptional activator NtrC1: structural studies of the regulatory and AAA+ ATPase domains. , 2003, Genes & development.

[16]  David J. Studholme,et al.  The Bacterial Enhancer-Dependent ς54(ςN) Transcription Factor , 2000, Journal of bacteriology.

[17]  N. Guex,et al.  SWISS‐MODEL and the Swiss‐Pdb Viewer: An environment for comparative protein modeling , 1997, Electrophoresis.

[18]  S. Kustu,et al.  The bacterial enhancer-binding protein NtrC as a molecular machine. , 1998, Cold Spring Harbor symposia on quantitative biology.

[19]  M. Buck,et al.  Mechanochemical ATPases and transcriptional activation , 2002, Molecular microbiology.

[20]  J. Cronan,et al.  Two-Carbon Compounds and Fatty Acids as Carbon Sources , 2005, EcoSal Plus.

[21]  A. J. Clifford,et al.  BIOCHIMICA ET BIOPHYSICA ACTA , 2022 .

[22]  W. Fong,et al.  The appearance of an ornithine decarboxylase inhibitory protein upon the addition of putrescine to cell cultures. , 1976, Biochimica et biophysica acta.

[23]  W. Delano.  The PyMOL Molecular Graphics System , 2002 .

[24]  W. D. Nunn,et al.  Genetic and molecular characterization of the genes involved in short-chain fatty acid degradation in Escherichia coli: the ato system , 1987, Journal of bacteriology.

[25]  R. Utsumi,et al.  Functional Characterization in Vitro of All Two-component Signal Transduction Systems from Escherichia coli* , 2005, Journal of Biological Chemistry.

[26]  A. Wedel,et al.  The bacterial enhancer-binding protein NTRC is a molecular machine: ATP hydrolysis is coupled to transcriptional activation. , 1995, Genes & development.

[27]  Ann M Stock,et al.  Histidine kinases and response regulator proteins in two-component signaling systems. , 2001, Trends in biochemical sciences.

[28]  S. Kustu,et al.  Constitutive forms of the enhancer-binding protein NtrC: evidence that essential oligomerization determinants lie in the central activation domain. , 1995, Journal of molecular biology.

[29]  B. Wanner,et al.  Phenotype MicroArray Analysis of Escherichia coli K-12 Mutants with Deletions of All Two-Component Systems , 2003, Journal of bacteriology.

[30]  W. Fong,et al.  Induction of a protein inhibitor to ornithine decarboxylase by the end products of its reaction. , 1976, Proceedings of the National Academy of Sciences of the United States of America.

[31]  M. M. Bradford.  A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. , 1976, Analytical biochemistry.

[32]  S. Austin,et al.  The prokaryotic enhancer binding protein NTRC has an ATPase activity which is phosphorylation and DNA dependent. , 1992, The EMBO journal.

[33]  T. N. Bhat,et al.  The Protein Data Bank , 2000, Nucleic Acids Res..

[34]  D. Kyriakidis,et al.  Regulation of the Escherichia coli biosynthetic ornithine decarboxylase activity by phosphorylation and nucleotides. , 1996, Biochimica et biophysica acta.

[35]  C. Panagiotidis,et al.  Physical and functional interactions between herpes simplex virus immediate-early proteins ICP4 and ICP27 , 1997, Journal of virology.

[36]  L. Hendershot,et al.  Characterization of the Nucleotide Binding Properties and ATPase Activity of Recombinant Hamster BiP Purified from Bacteria (*) , 1995, The Journal of Biological Chemistry.

[37]  T. Mizuno,et al.  Compilation of all genes encoding two-component phosphotransfer signal transducers in the genome of Escherichia coli. , 1997, DNA research : an international journal for rapid publication of reports on genes and genomes.

[38]  M. Marzioch,et al.  Two complementary approaches to study peroxisome biogenesis in Saccharomyces cerevisiae: forward and reversed genetics. , 1993, Biochimie.

[39]  F. Neidhardt,et al.  Escherichia Coli and Salmonella: Typhimurium Cellular and Molecular Biology , 1987 .

[40]  Rolf Apweiler,et al.  The SWISS-PROT protein sequence database and its supplement TrEMBL in 2000 , 2000, Nucleic Acids Res..

[41]  J. Stock,et al.  Phosphorylation of bacterial response regulator proteins by low molecular weight phospho-donors. , 1992, Proceedings of the National Academy of Sciences of the United States of America.

[42]  P. Overath,et al.  ato Operon: a highly inducible system for acetoacetate and butyrate degradation in Escherichia coli. , 1972, European journal of biochemistry.

[43]  R. Dixon,et al.  Domain architectures of sigma54-dependent transcriptional activators. , 2003, Journal of bacteriology.

[44]  S. Kustu,et al.  The phosphorylated form of the enhancer-binding protein NTRC has an ATPase activity that is essential for activation of transcription , 1991, Cell.

[45]  D. Kyriakidis,et al.  Identification, cloning, and nucleotide sequencing of the ornithine decarboxylase antizyme gene of Escherichia coli. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[46]  Takeshi Mizuno,et al.  Transcriptome analysis of all two‐component regulatory system mutants of Escherichia coli K‐12 , 2002, Molecular microbiology.

[47]  Alan E. Mark,et al.  The GROMOS96 Manual and User Guide , 1996 .

[48]  John B. Anderson,et al.  CDD: a curated Entrez database of conserved domain alignments , 2003, Nucleic Acids Res..

[49]  E. Lioliou,et al.  The role of bacterial antizyme: From an inhibitory protein to AtoC transcriptional regulator , 2004, Microbial cell factories.

[50]  J. Thornton,et al.  PROCHECK: a program to check the stereochemical quality of protein structures , 1993 .

[51]  B. T. Nixon,et al.  Constitutive ATP hydrolysis and transcription activation by a stable, truncated form of Rhizobium meliloti DCTD, a sigma 54-dependent transcriptional activator. , 1994, The Journal of biological chemistry.

[52]  Ann M Stock,et al.  Two-component signal transduction. , 2000, Annual review of biochemistry.

[53]  Yan Zhu,et al.  Solution structure of the homodimeric core domain of Escherichia coli histidine kinase EnvZ , 1999, Nature Structural Biology.

[54]  S. Austin,et al.  Properties of a mutant form of the prokaryotic enhancer binding protein, NTRC, which hydrolyses ATP in the absence of effectors , 1998, FEBS letters.