Structure of the pilus assembly protein TadZ from Eubacterium rectale: implications for polar localization

The tad (tight adherence) locus encodes a protein translocation system that produces a novel variant of type IV pili. The pilus assembly protein TadZ (called CpaE in Caulobacter crescentus) is ubiquitous in tad loci, but is absent in other type IV pilus biogenesis systems. The crystal structure of TadZ from Eubacterium rectale (ErTadZ), in complex with ATP and Mg2+, was determined to 2.1 Å resolution. ErTadZ contains an atypical ATPase domain with a variant of a deviant Walker‐A motif that retains ATP binding capacity while displaying only low intrinsic ATPase activity. The bound ATP plays an important role in dimerization of ErTadZ. The N‐terminal atypical receiver domain resembles the canonical receiver domain of response regulators, but has a degenerate, stripped‐down ‘active site’. Homology modelling of the N‐terminal atypical receiver domain of CpaE indicates that it has a conserved protein–protein binding surface similar to that of the polar localization module of the social mobility protein FrzS, suggesting a similar function. Our structural results also suggest that TadZ localizes to the pole through the atypical receiver domain during an early stage of pili biogenesis, and functions as a hub for recruiting other pili components, thus providing insights into the Tad pilus assembly process.

[1]  Mitchell D. Miller,et al.  A conserved fold for fimbrial components revealed by the crystal structure of a putative fimbrial assembly protein (BT1062) from Bacteroides thetaiotaomicron at 2.2 Å resolution , 2010, Acta crystallographica. Section F, Structural biology and crystallization communications.

[2]  B. Kazmierczak,et al.  Analysis of FimX, a phosphodiesterase that governs twitching motility in Pseudomonas aeruginosa , 2006, Molecular microbiology.

[3]  R. DeSalle,et al.  flp‐1, the first representative of a new pilin gene subfamily, is required for non‐specific adherence of Actinobacillus actinomycetemcomitans , 2001, Molecular microbiology.

[4]  E. Baker,et al.  Pili in Gram-negative and Gram-positive bacteria — structure, assembly and their role in disease , 2009, Cellular and Molecular Life Sciences.

[5]  Aina E Cohen,et al.  An automated system to mount cryo-cooled protein crystals on a synchrotron beam line, using compact sample cassettes and a small-scale robot. , 2002, Journal of applied crystallography.

[6]  E. Huitema,et al.  Bacterial Birth Scar Proteins Mark Future Flagellum Assembly Site , 2006, Cell.

[7]  Yang Zhang,et al.  I-TASSER server for protein 3D structure prediction , 2008, BMC Bioinformatics.

[8]  M. Tomich,et al.  The tad locus: postcards from the widespread colonization island , 2007, Nature Reviews Microbiology.

[9]  S. Golden,et al.  The pseudo‐receiver domain of CikA regulates the cyanobacterial circadian input pathway , 2006, Molecular microbiology.

[10]  E. O. Jordan,et al.  Bacterial Motility. , 1934, Journal of bacteriology.

[11]  Lucy Shapiro,et al.  A dynamically localized histidine kinase controls the asymmetric distribution of polar pili proteins , 2002, The EMBO journal.

[12]  H. Lam,et al.  The asymmetric spatial distribution of bacterial signal transduction proteins coordinates cell cycle events. , 2003, Developmental cell.

[13]  Nathan J Hillson,et al.  High-throughput identification of protein localization dependency networks , 2010, Proceedings of the National Academy of Sciences.

[14]  D. Fine,et al.  Tight-adherence genes of Actinobacillus actinomycetemcomitans are required for virulence in a rat model , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[15]  R. Bourret Receiver domain structure and function in response regulator proteins. , 2010, Current opinion in microbiology.

[16]  S. Kustu,et al.  Glutamate at the site of phosphorylation of nitrogen-regulatory protein NTRC mimics aspartyl-phosphate and activates the protein. , 1993, Journal of molecular biology.

[17]  M. Saier,et al.  Type II protein secretion and its relationship to bacterial type IV pili and archaeal flagella. , 2003, Microbiology.

[18]  Social motility in Myxococcus xanthus requires FrzS, a protein with an extensive coiled‐coil domain , 2000, Molecular microbiology.

[19]  H. Sondermann,et al.  Vibrio cholerae VpsT Regulates Matrix Production and Motility by Directly Sensing Cyclic di-GMP , 2010, Science.

[20]  David C. Richardson,et al.  MOLPROBITY: structure validation and all-atom contact analysis for nucleic acids and their complexes , 2004, Nucleic Acids Res..

[21]  H. Flint,et al.  Proposal of a neotype strain (A1-86) for Eubacterium rectale. Request for an opinion. , 2008, International journal of systematic and evolutionary microbiology.

[22]  Richard J Morris,et al.  Towards complete validated models in the next generation of ARP/wARP. , 2004, Acta crystallographica. Section D, Biological crystallography.

[23]  G. Crooks,et al.  WebLogo: a sequence logo generator. , 2004, Genome research.

[24]  T. Mizuno,et al.  Genes encoding pseudo-response regulators: insight into His-to-Asp phosphorelay and circadian rhythm in Arabidopsis thaliana. , 2000, Plant & cell physiology.

[25]  Kevin Cowtan,et al.  research papers Acta Crystallographica Section D Biological , 2005 .

[26]  J. Lutkenhaus,et al.  MinD and role of the deviant Walker A motif, dimerization and membrane binding in oscillation , 2003, Molecular microbiology.

[27]  Lucy Shapiro,et al.  A membrane metalloprotease participates in the sequential degradation of a Caulobacter polarity determinant , 2004, Molecular microbiology.

[28]  H. Lam,et al.  A Landmark Protein Essential for Establishing and Perpetuating the Polarity of a Bacterial Cell , 2006, Cell.

[29]  E. Birney,et al.  Pfam: the protein families database , 2013, Nucleic Acids Res..

[30]  A. Filloux,et al.  FppA, a Novel Pseudomonas aeruginosa Prepilin Peptidase Involved in Assembly of Type IVb Pili , 2006, Journal of bacteriology.

[31]  L. Shapiro,et al.  Identification of a localization factor for the polar positioning of bacterial structural and regulatory proteins , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[32]  L. Søgaard-Andersen,et al.  Coupling of protein localization and cell movements by a dynamically localized response regulator in Myxococcus xanthus , 2007, The EMBO journal.

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

[34]  D. Kaiser,et al.  Type IV pili and cell motility , 1999, Molecular microbiology.

[35]  Earl W. Cornell,et al.  An approach to rapid protein crystallization using nanodroplets , 2002 .

[36]  C. Sáez,et al.  Analysis of MinD Mutations Reveals Residues Required for MinE Stimulation of the MinD ATPase and Residues Required for MinC Interaction , 2005, Journal of bacteriology.

[37]  Herbert L. Axelrod,et al.  Structural Biology and Crystallization Communications Structure of a Membrane-attack Complex/perforin (macpf) Family Protein from the Human Gut Symbiont Bacteroides Thetaiotaomicron , 2022 .

[38]  R. J. Gorlin Asymmetry. , 1887, American journal of medical genetics.

[39]  Patrick T McGrath,et al.  A phospho-signaling pathway controls the localization and activity of a protease complex critical for bacterial cell cycle progression. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[40]  K. Morikawa,et al.  Structural and functional studies of MinD ATPase: implications for the molecular recognition of the bacterial cell division apparatus , 2001, The EMBO journal.

[41]  L. Shapiro,et al.  MipZ, a Spatial Regulator Coordinating Chromosome Segregation with Cell Division in Caulobacter , 2006, Cell.

[42]  Jeffrey M. Skerker,et al.  Identification and cell cycle control of a novel pilus system in Caulobacter crescentus , 2000, The EMBO journal.

[43]  T. Leonard,et al.  Bacterial chromosome segregation: structure and DNA binding of the Soj dimer — a conserved biological switch , 2005, The EMBO journal.

[44]  P. Evans,et al.  Scaling and assessment of data quality. , 2006, Acta crystallographica. Section D, Biological crystallography.

[45]  Thomas R Ioerger,et al.  Crystal Structure of Circadian Clock Protein KaiA from Synechococcus elongatus* , 2004, Journal of Biological Chemistry.

[46]  R. DeSalle,et al.  Nonspecific Adherence by Actinobacillus actinomycetemcomitans Requires Genes Widespread inBacteria and Archaea , 2000, Journal of bacteriology.

[47]  Eric Koesema,et al.  Combining the polymerase incomplete primer extension method for cloning and mutagenesis with microscreening to accelerate structural genomics efforts , 2008, Proteins.

[48]  D. Wemmer,et al.  An atypical receiver domain controls the dynamic polar localization of the Myxococcus xanthus social motility protein FrzS , 2007, Molecular microbiology.

[49]  T. Mizuno,et al.  Pseudo-Response Regulators (PRRs) or True Oscillator Components (TOCs). , 2005, Plant & cell physiology.

[50]  D. Fine,et al.  Nonspecific Adherence and Fibril Biogenesis by Actinobacillus actinomycetemcomitans: TadA Protein Is an ATPase , 2001, Journal of bacteriology.

[51]  Adam Godzik,et al.  Structural genomics of the Thermotoga maritima proteome implemented in a high-throughput structure determination pipeline , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[52]  Y. Brun,et al.  A Molecular Beacon Defines Bacterial Cell Asymmetry , 2006, Cell.

[53]  Kurt Wüthrich,et al.  Structural Biology and Crystallization Communications the Jcsg High-throughput Structural Biology Pipeline , 2022 .

[54]  E V Koonin,et al.  A superfamily of ATPases with diverse functions containing either classical or deviant ATP-binding motif. , 1993, Journal of molecular biology.

[55]  J. Lutkenhaus,et al.  Dynamic assembly of MinD on phospholipid vesicles regulated by ATP and MinE , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[56]  T. Mignot,et al.  Two localization motifs mediate polar residence of FrzS during cell movement and reversals of Myxococcus xanthus , 2007, Molecular microbiology.

[57]  S. Golden,et al.  Structure and function from the circadian clock protein KaiA of Synechococcus elongatus: A potential clock input mechanism , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[58]  T. Mignot,et al.  Bacterial motility complexes require the actin‐like protein, MreB and the Ras homologue, MglA , 2010, The EMBO journal.

[59]  G. Mahairas,et al.  Haemophilus ducreyi Requires the flp Gene Cluster for Microcolony Formation In Vitro , 2002, Infection and Immunity.

[60]  B. Giese,et al.  Cell cycle-dependent dynamic localization of a bacterial response regulator with a novel di-guanylate cyclase output domain. , 2004, Genes & development.

[61]  L. Holm,et al.  The Pfam protein families database , 2005, Nucleic Acids Res..

[62]  R. DeSalle,et al.  Genes for tight adherence of Actinobacillus actinomycetemcomitans: from plaque to plague to pond scum. , 2001, Trends in microbiology.

[63]  L. Rothfield,et al.  The MinD protein is a membrane ATPase required for the correct placement of the Escherichia coli division site. , 1991, The EMBO journal.

[64]  B. Giese,et al.  Structural basis of activity and allosteric control of diguanylate cyclase. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[65]  G Bricogne,et al.  Generation, representation and flow of phase information in structure determination: recent developments in and around SHARP 2.0. , 2003, Acta crystallographica. Section D, Biological crystallography.

[66]  S. Golden,et al.  NMR structure of the pseudo‐receiver domain of CikA , 2007, Protein Science.

[67]  Rob DeSalle,et al.  The Widespread Colonization Island of Actinobacillus actinomycetemcomitans , 2003, Nature Genetics.

[68]  G. Sheldrick A short history of SHELX. , 2008, Acta crystallographica. Section A, Foundations of crystallography.

[69]  G. Petsko,et al.  Structure of the Mg(2+)-bound form of CheY and mechanism of phosphoryl transfer in bacterial chemotaxis. , 1994, Biochemistry.

[70]  J. Jonasson Haemophilus Ducreyi , 1993, International journal of STD & AIDS.

[71]  D. Figurski,et al.  The product of tadZ, a new member of the parA/minD superfamily, localizes to a pole in Aggregatibacter actinomycetemcomitans , 2012, Molecular microbiology.

[72]  Stephen K Burley,et al.  Structural genomics , 1999, Current Biology.