Structure of the periplasmic domain of Pseudomonas aeruginosa TolA: evidence for an evolutionary relationship with the TonB transporter protein

The crystal structure of the C‐terminal domain III of Pseudomonas aeruginosa TolA has been determined at 1.9 Å resolution. The fold is similar to that of the corresponding domain of Escherichia coli TolA, despite the limited amino acid sequence identity of the two proteins (20%). A pattern was discerned that conserves the fold of domain III within the wider TolA family and, moreover, reveals a relationship between TolA domain III and the C‐terminal domain of the TonB transporter proteins. We propose that the TolA and TonB C‐terminal domains have a common evolutionary origin and are related by means of domain swapping, with interesting mechanistic implications. We have also determined the overall shape of the didomain, domains II + III, of P.aeruginosa TolA by solution X‐ray scattering. The molecule is monomeric—its elongated, stalk shape can accommodate the crystal structure of domain III at one end, and an elongated helical bundle within the portion corresponding to domain II. Based on these data, a model for the periplasmic domains of P.aeruginosa TolA is presented that may explain the inferred allosteric properties of members of the TolA family. The mechanisms of TolA‐mediated entry of bateriophages in P.aeruginosa and E.coli are likely to be similar.

[1]  G. Porod,et al.  Die Röntgenkleinwinkelstreuung von dichtgepackten kolloiden Systemen , 1952 .

[2]  R. Lewis Multiwire Gas Proportional Counters: Decrepit Antiques or ClassicPerformers? , 1994, Journal of synchrotron radiation.

[3]  G. Moeck,et al.  Characterization of In Vitro Interactions between a Truncated TonB Protein fromEscherichia coli and the Outer Membrane Receptors FhuA and FepA , 2001, Journal of bacteriology.

[4]  R. Lewis,et al.  Time‐resolved x‐ray diffraction station: X‐ray optics, detectors, and data acquisition , 1989 .

[5]  R. E. Webster,et al.  Nucleotide sequence of a gene cluster involved in entry of E colicins and single-stranded DNA of infecting filamentous bacteriophages into Escherichia coli , 1987, Journal of bacteriology.

[6]  C. Lazdunski,et al.  Colicin import and pore formation: a system for studying protein transport across membranes? , 1995, Molecular microbiology.

[7]  R. Lloubès,et al.  Binding of colicins A and El to purified TolA domains. , 1997, Microbiology.

[8]  H. Nikaido,et al.  Porins and specific channels of bacterial outer membranes , 1992, Molecular microbiology.

[9]  A Wlodawer,et al.  Filamentous phage infection: crystal structure of g3p in complex with its coreceptor, the C-terminal domain of TolA. , 1999, Structure.

[10]  J. Lazzaroni,et al.  Mutational Analysis of the Escherichia coli K-12 TolA N-Terminal Region and Characterization of Its TolQ-Interacting Domain by Genetic Suppression , 1998, Journal of bacteriology.

[11]  M. Gavioli,et al.  Proton motive force drives the interaction of the inner membrane TolA and outer membrane Pal proteins in Escherichia coli , 2000, Molecular microbiology.

[12]  J. Lazzaroni,et al.  Protein complex within Escherichia coli inner membrane. TolA N-terminal domain interacts with TolQ and TolR proteins , 1995, The Journal of Biological Chemistry.

[13]  E. Bouveret,et al.  Colicin Import into Escherichia coli Cells , 1998, Journal of bacteriology.

[14]  R. E. Webster,et al.  The TolA protein interacts with colicin E1 differently than with other group A colicins , 1997, Journal of bacteriology.

[15]  G. Murshudov,et al.  Refinement of macromolecular structures by the maximum-likelihood method. , 1997, Acta crystallographica. Section D, Biological crystallography.

[16]  T L Blundell,et al.  FUGUE: sequence-structure homology recognition using environment-specific substitution tables and structure-dependent gap penalties. , 2001, Journal of molecular biology.

[17]  A. V. Semenyuk,et al.  GNOM – a program package for small-angle scattering data processing , 1991 .

[18]  T. Blundell,et al.  Definition of general topological equivalence in protein structures. A procedure involving comparison of properties and relationships through simulated annealing and dynamic programming. , 1990, Journal of molecular biology.

[19]  J. Lakey,et al.  Discovery of critical Tol A‐binding residues in the bactericidal toxin colicin N: a biophysical approach , 1998, Molecular microbiology.

[20]  S. Grzesiek,et al.  NMRPipe: A multidimensional spectral processing system based on UNIX pipes , 1995, Journal of biomolecular NMR.

[21]  Andrew E. Torda,et al.  Sausage: protein threading with flexible force fields , 1999, Bioinform..

[22]  R. E. Webster,et al.  TolA: a membrane protein involved in colicin uptake contains an extended helical region. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[23]  M. Valvano,et al.  Surface expression of O‐specific lipopolysaccharide in Escherichia coli requires the function of the TolA protein , 2000, Molecular microbiology.

[24]  V. Braun,et al.  Evolutionary relationship of uptake systems for biopolymers in Escherichia coli: cross‐complementation between the TonB‐ExbB‐ExbD and the TolA‐TolQ‐TolR proteins , 1993, Molecular microbiology.

[25]  J. Ramos,et al.  Mutations in Each of the tol Genes ofPseudomonas putida Reveal that They Are Critical for Maintenance of Outer Membrane Stability , 2000, Journal of bacteriology.

[26]  J. Lakey,et al.  The TolA-recognition site of colicin N. ITC, SPR and stopped-flow fluorescence define a crucial 27-residue segment. , 2000, Journal of molecular biology.

[27]  G. Porod Die Röntgenkleinwinkelstreuung von dichtgepackten kolloiden Systemen , 1953 .

[28]  E. Pai,et al.  The structure of enzyme IIAlactose from Lactococcus lactis reveals a new fold and points to possible interactions of a multicomponent system. , 1997, Structure.

[29]  S. Luria,et al.  Genetics and Physiology of Colicin-tolerant Mutants of Escherichia coli , 1967, Journal of bacteriology.

[30]  M. Nomura,et al.  Interaction of Colicins with Bacterial Cells III. Colicin-tolerant Mutations in Escherichia coli , 1967, Journal of bacteriology.

[31]  E. R. Lafontaine,et al.  Identification and characterization of the tolQRA genes of Pseudomonas aeruginosa , 1996, Journal of bacteriology.

[32]  Dmitri I. Svergun,et al.  Automated matching of high- and low-resolution structural models , 2001 .

[33]  V. Braun,et al.  Import of biopolymers into Escherichia coli: nucleotide sequences of the exbB and exbD genes are homologous to those of the tolQ and tolR genes, respectively , 1989, Journal of bacteriology.

[34]  J. Konisky,et al.  Interaction of colicins with bacterial cells. II. Specific alteration of Escherichia coli ribosomes induced by colicin E3 in vivo. , 1967, Journal of molecular biology.

[35]  E. Bouveret,et al.  Import of colicins across the outer membrane of Escherichia coli involves multiple protein interactions in the periplasm , 2001, Molecular microbiology.

[36]  R. E. Webster,et al.  fii, a bacterial locus required for filamentous phage infection and its relation to colicin-tolerant tolA and tolB , 1986, Journal of bacteriology.

[37]  C. Calladine,et al.  How to untwist an alpha-helix: structural principles of an alpha-helical barrel. , 2001, Journal of molecular biology.

[38]  R M Esnouf,et al.  Further additions to MolScript version 1.4, including reading and contouring of electron-density maps. , 1999, Acta crystallographica. Section D, Biological crystallography.

[39]  B. Berger,et al.  MultiCoil: A program for predicting two‐and three‐stranded coiled coils , 1997, Protein science : a publication of the Protein Society.

[40]  J. Deisenhofer,et al.  Structural basis of gating by the outer membrane transporter FecA. , 2002, Science.

[41]  R. E. Webster,et al.  Filamentous phage infection: required interactions with the TolA protein , 1997, Journal of bacteriology.

[42]  L. Riechmann,et al.  The C-Terminal Domain of TolA Is the Coreceptor for Filamentous Phage Infection of E. coli , 1997, Cell.

[43]  Charlotte M. Deane,et al.  JOY: protein sequence-structure representation and analysis , 1998, Bioinform..

[44]  A. Wlodawer,et al.  Crystal Structure of the Dimeric C-terminal Domain of TonB Reveals a Novel Fold* , 2001, The Journal of Biological Chemistry.

[45]  F. Crick,et al.  The packing of α‐helices: simple coiled‐coils , 1953 .

[46]  D. Vos,et al.  Molecular and immunological characterization of OprL, the 18 kDa outer-membrane peptidoglycan-associated lipoprotein (PAL) of Pseudomonas aeruginosa. , 1997, Microbiology.

[47]  D I Svergun,et al.  Determination of domain structure of proteins from X-ray solution scattering. , 2001, Biophysical journal.

[48]  P Argos,et al.  Intramolecular cavities in globular proteins. , 1994, Protein engineering.

[49]  A. Lupas,et al.  Predicting coiled-coil regions in proteins. , 1997, Current opinion in structural biology.

[50]  J. Sturgis,et al.  Organisation and evolution of the tol-pal gene cluster. , 2001, Journal of molecular microbiology and biotechnology.

[51]  Z. Otwinowski,et al.  Processing of X-ray diffraction data collected in oscillation mode. , 1997, Methods in enzymology.

[52]  I. Holland,et al.  Genetic Basis of Colicin E Susceptibility in Escherichia coli I. Isolation and Properties of Refractory Mutants and the Preliminary Mapping of Their Mutations , 1967, Journal of bacteriology.

[53]  C. Higgins,et al.  A sequence‐specific function for the N‐terminal signal‐like sequence of the TonB protein , 1993, Molecular microbiology.

[54]  D. Svergun,et al.  CRYSOL : a program to evaluate X-ray solution scattering of biological macromolecules from atomic coordinates , 1995 .

[55]  J. Sturgis,et al.  The TolQ–TolR proteins energize TolA and share homologies with the flagellar motor proteins 
MotA–MotB , 2001, Molecular microbiology.

[56]  J. Lazzaroni,et al.  Energy-Dependent Conformational Change in the TolA Protein ofEscherichia coli Involves Its N-Terminal Domain, TolQ, and TolR , 2001, Journal of bacteriology.

[57]  P. Kraulis A program to produce both detailed and schematic plots of protein structures , 1991 .

[58]  G. Barton,et al.  Multiple protein sequence alignment from tertiary structure comparison: Assignment of global and residue confidence levels , 1992, Proteins.

[59]  S. Sasso,et al.  Circular dichroism and molecular modeling of the E. coli TolA periplasmic domains. , 1999, Biospectroscopy.

[60]  George M Sheldrick,et al.  [37] Patterson superposition and ab initio phasing. , 1997, Methods in enzymology.

[61]  Collaborative Computational,et al.  The CCP4 suite: programs for protein crystallography. , 1994, Acta crystallographica. Section D, Biological crystallography.

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

[63]  J. Popot,et al.  Transmembrane alpha-helix interactions are required for the functional assembly of the Escherichia coli Tol complex. , 1995, Journal of molecular biology.

[64]  S. Zakharov,et al.  Structural stability and domain organization of colicin E1. , 2000, Journal of molecular biology.

[65]  D Eisenberg,et al.  3D domain swapping: A mechanism for oligomer assembly , 1995, Protein science : a publication of the Protein Society.

[66]  K H Kim,et al.  Deciphering the design of the tropomyosin molecule , 2001, Proceedings of the National Academy of Sciences of the United States of America.

[67]  E A Merritt,et al.  Raster3D Version 2.0. A program for photorealistic molecular graphics. , 1994, Acta crystallographica. Section D, Biological crystallography.

[68]  C. Boulin,et al.  Data appraisal, evaluation and display for synchrotron radiation experiments: Hardware and software , 1986 .

[69]  R. Kadner,et al.  Sequence Changes in the Ton Box Region of BtuB Affect Its Transport Activities and Interaction with TonB Protein , 2000, Journal of bacteriology.

[70]  Alfonso Mondragón,et al.  Structures of Two Repeats of Spectrin Suggest Models of Flexibility , 1999, Cell.

[71]  D. I. Svergun,et al.  Structure Analysis by Small-Angle X-Ray and Neutron Scattering , 1987 .

[72]  R. Lloubès,et al.  Escherichia coli tol-pal Mutants Form Outer Membrane Vesicles , 1998, Journal of bacteriology.

[73]  R F Standaert,et al.  Atomic structures of the human immunophilin FKBP-12 complexes with FK506 and rapamycin. , 1993, Journal of molecular biology.