Teresa Ruiz

University of Vermont, Burlington, Vermont, United States

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Publications (69)179.87 Total impact

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    ABSTRACT: Morphogenesis protein C (MorC) of Aggregatibacter actinomycetemcomitans is important for maintaining the membrane morphology and integrity of the cell envelope of this oral pathogen. The MorC sequence and operon organization was found to be conserved in Gammaproteobacteria, based on a bioinformatic analysis of 435 sequences from representative organisms. Functional conservation of MorC was investigated utilizing an A. actinomycetemcomitans morC mutant as a model system to express MorC homologs from four phylogenetically diverse representatives of the Gammaproteobacteria: Haemophilus influenzae; Escherichia coli; Pseudomonas aeruginosa; and Moraxella catarrhalis. The A. actinomycetemcomitans strains expressing the homologous proteins were assessed for sensitivity to bile salts, leukotoxin secretion, autoaggregation and membrane morphology. MorC from the most closely related organism (H. influenzae) was functionally identical to MorC from A. actinomycetemcomitans. However, the genes from more distantly related organisms restored some but not all A. actinomycetemcomitans mutant phenotypes. In addition, deletion mutagenesis indicated that the most conserved portion of the protein, the carboxyl terminus DUF490 domain, was necessary to maintain the integrity of the membrane. Deletion of the last ten amino acids of this domain of the A. actinomycetemcomitans MorC protein was sufficient to disrupt membrane stability and leukotoxin secretion. The data suggest that the MorC sequence is functionally conserved across Gammaproteobacteria and the carboxyl terminus of the protein is essential for maintaining membrane physiology. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    07/2015; DOI:10.1111/omi.12120

  • Microscopy and Microanalysis 08/2014; 20(S3):1190-1191. DOI:10.1017/S1431927614007685 · 1.88 Impact Factor
  • Shaun Benjamin · Michael Radermacher · Teresa Ruiz ·

    Microscopy and Microanalysis 08/2014; 20(S3):1246-1247. DOI:10.1017/S143192761400796X · 1.88 Impact Factor
  • K. SMITH · R. VOOGT · T. RUIZ · K. MINTZ ·
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    ABSTRACT: Outer membranes (OM) of Gram-negative bacteria contain proteins important for pathogenesis: adhesins; invasins; transporters; and toxin secretion machineries. OMs can be classified according to membrane morphology: flat or convoluted/rugose. The basis and biological significance for this difference are understudied. Aggregatibacter actinomycetemcomitans morC, coding for a 141 kDa membrane protein, when inactivated changes the OM from rugose to flat. The mutant is also defective in leukotoxin secretion and displays sensitivity to bile salts, a membrane destabilizing agent. Objective: To determine if transformation of the A. actinomycetemcomitans mutant with morC from organisms with flat OMs complement the phenotypes. Methods: Selection of morC genes was determined based on membrane morphology and bioinformatic analysis of sequence homology and operon organization from seven families of Gammaproteobacteria. Three representative homologs of morC were amplified by PCR and transformed into the A. actinomycetemcomitans mutant on a replicating plasmid. The resulting strains were assayed for complementation of: leukotoxin secretion by immunoblot; membrane stability by growth in bile salts; and membrane morphology by transmission electron microscopy. Immunoblots was used to determine MorC stability. Results: Bioinformatic analyses demonstrated that the MorC sequence and operon organization are conserved. Complementation experiments revealed that H. influenzae morC, from the same bacterial family expressing a rugose OM, and a homologous protein from E. coli, expressing a flat membrane morphology, restored MorC function. However, a distantly related bacterium (Pseudomonas aeruginosa) high in G/C nucleotide content expressing a flat membrane morphology, did not complement. Conclusions: morC is conserved in the genome of over 500 bacteria independent of the membrane morphology. Interestingly, the complementation of the morC mutant by E. coli morC implies that MorC may not be directly associated with membrane morphology in A. actinomycetemcomitans, but acts as a structural protein to nucleate proteins associated with this phenotype.
    IADR/AADR/CADR General Session and Exhibition 2013; 03/2013
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    ABSTRACT: Serine-rich repeat glycoproteins (SRRPs) are important bacterial adhesins conserved in streptococci and staphylococci. Fap1, a SRRP identified in Streptococcus parasanguinis, is the major constituent of bacterial fimbriae and is required for adhesion and biofilm formation. An 11-gene cluster is required for Fap1 glycosylation and secretion; however, the exact mechanism of Fap1 biogenesis remains a mystery. Two glycosylation-associated proteins within this cluster—Gap1 and Gap3—function together in Fap1 biogenesis. Here we report the role of the third glycosylation-associated protein, Gap2. A gap2 mutant exhibited the same phenotype as the gap1 and gap3 mutants in terms of Fap1 biogenesis, fimbrial assembly, and bacterial adhesion, suggesting that the three proteins interact. Indeed, all three proteins interacted with each other independently and together to form a stable protein complex. Mechanistically, Gap2 protected Gap3 from degradation by ClpP protease, and Gap2 required the presence of Gap1 for expression at the wild-type level. Gap2 augmented the function of Gap1 in stabilizing Gap3; this function was conserved in Gap homologs from Streptococcus agalactiae. Our studies demonstrate that the three Gap proteins work in concert in Fap1 biogenesis and reveal a new function of Gap2. This insight will help us elucidate the molecular mechanism of SRRP biogenesis in this bacterium and in pathogenic species.
    Journal of bacteriology 03/2013; 195(10). DOI:10.1128/JB.02255-12 · 2.81 Impact Factor
  • Lingbo Yu · Robert R Snapp · Teresa Ruiz · Michael Radermacher ·
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    ABSTRACT: When heterogeneous samples of macromolecular assemblies are being examined by 3D electron microscopy (3DEM), often multiple reconstructions are obtained. For example, subtomograms of individual particles can be acquired from tomography, or volumes of multiple 2D classes can be obtained by random conical tilt reconstruction. Of these, similar volumes can be averaged to achieve higher resolution. Volume alignment is an essential step before 3D classification and averaging. Here we present a projection-based volume alignment (PBVA) algorithm. We select a set of projections to represent the reference volume and align them to a second volume. Projection alignment is achieved by maximizing the cross-correlation function with respect to rotation and translation parameters. If data are missing, the cross-correlation functions are normalized accordingly. Accurate alignments are obtained by averaging and quadratic interpolation of the cross-correlation maximum. Comparisons of the computation time between PBVA and traditional 3D cross-correlation methods demonstrate that PBVA outperforms the traditional methods. Performance tests were carried out with different signal-to-noise ratios using modeled noise and with different percentages of missing data using a cryo-EM dataset. All tests show that the algorithm is robust and highly accurate. PBVA was applied to align the reconstructions of a subcomplex of the NADH: ubiquinone oxidoreductase (Complex I) from the yeast Yarrowia lipolytica, followed by classification and averaging.
    Journal of Structural Biology 02/2013; 182(2). DOI:10.1016/j.jsb.2013.01.011 · 3.23 Impact Factor
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    ABSTRACT: Bacterial membranes serve as selective environmental barriers and contain determinants required for bacterial colonization and survival. Cell envelopes of Gram-negative bacteria consist of an outer and an inner membrane separated by a periplasmic space. Most Gram-negative bacteria display a smooth outer surface (e.g. Enterobacteriaceae), whereas members of the Pasteurellaceae and Moraxellaceae families show convoluted surfaces. Aggregatibacter actinomycetemcomitans, an oral pathogen representative of the Pasteurellaceae family, displays a convoluted membrane morphology. This phenotype is associated with the presence of morphogenesis protein C (MorC). Inactivation of the morC gene results in a smooth membrane appearance when visualized by 2D electron microscopy. In this study, 3D electron microscopy and atomic force microscopy of whole-mount bacterial preparations as well as 3D electron microscopy of ultra-thin sections of high-pressure frozen and freeze-substituted specimens were employed to characterize the membranes of both wild type and morC mutant strains of A. actinomycetemcomitans. Our results show that the mutant strain contains fewer convolutions than the wild-type bacterium, which exhibits a higher curvature of the outer membrane and a periplasmic space with two fold larger volume/area ratio than the mutant bacterium. The inner membrane of both strains has a smooth appearance and shows connections with the outer membrane, as revealed by visualization and segmentation of 3D tomograms. The present studies and the availability of genetically modified organisms with altered outer membrane morphology make A. actinomycetemcomitans a model organism for examining membrane remodeling and its implications in antibiotic resistance and virulence in the Pasteurellaceae and Moraxellaceae bacterial families.
    Journal of bacteriology 02/2013; DOI:10.1128/JB.02149-12 · 2.81 Impact Factor
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    X Jiang · T Ruiz · K P Mintz ·
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    ABSTRACT: The extracellular matrix protein adhesin A (EmaA) surface antennae-like structures of the periodontal pathogen Aggregatibacter actinomycetemcomitans are composed of three identical protein monomers. Recently, we have demonstrated that the protein is synthesized with an extended signal peptide of 56 amino acids necessary for membrane targeting and protein translocation. In this study, EmaA secretion was demonstrated to be reliant on a chaperone-dependent secretion pathway. Deletion of secB partially reduced but did not abolish the amount of EmaA in the membrane. This observation was attributed to an increase in the synthesis of DnaK in the ΔsecB strain. Overexpression of a DnaK substitution mutant (A174T), with diminished activity, in the ΔsecB strain further reduced the amount of EmaA in the membrane. Expression of dnaK A174T in the wild-type strain did not affect the amount of EmaA in the membrane when grown under optimal growth conditions at 37°C. However, EmaA was found to be reduced when this strain was grown at heat-shock temperature. A chromosomal deletion of amino acids 16-39 of the EmaA extended signal peptide, transformed with either the wild-type or dnaK A174T-expressing plasmid, did not affect the amount of EmaA in the membrane. In addition, the level of EmaA in a ΔsecB/emaA(-) double mutant strain expressing EmaAΔ16-39 was unchanged when grown at both temperatures. The data suggest that chaperones are required for the targeting of EmaA to the membrane and a specific region of the signal peptide is necessary for secretion under stress conditions.
    10/2012; 27(5):382-96. DOI:10.1111/j.2041-1014.2012.00652.x
  • L. R. Nyland · M. Radermacher · K. Mintz · T. Ruiz ·

    Microscopy and Microanalysis 07/2012; 18(S2):86-87. DOI:10.1017/S1431927612002280 · 1.88 Impact Factor

  • Microscopy and Microanalysis 07/2012; 18(S2):88-89. DOI:10.1017/S1431927612002292 · 1.88 Impact Factor
  • Y. Zhang · T. Ruiz · M. Radermacher ·

    Microscopy and Microanalysis 07/2012; 18(S2):122-123. DOI:10.1017/S1431927612002462 · 1.88 Impact Factor
  • M. Radermacher · L. Yu · F. Azari · L. Nyland · T. Ruiz ·

    Microscopy and Microanalysis 07/2012; 18(S2):118-119. DOI:10.1017/S1431927612002449 · 1.88 Impact Factor
  • L Yu · R R Snapp · T Ruiz · M Radermacher ·
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    ABSTRACT: Extended abstract of a paper presented at Microscopy and Microanalysis 2012 in Phoenix, Arizona, USA, July 29 - August 2, 2012.
    Microscopy and Microanalysis 07/2012; 18 Suppl 2:66-7. DOI:10.1017/S1431927612002188 · 1.88 Impact Factor
  • F. Azari · L. Nyland · T. Ruiz · C. Yu · K. Mintz ·

    Microscopy and Microanalysis 07/2012; 18(S2):82-83. DOI:10.1017/S1431927612002267 · 1.88 Impact Factor
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    Gaoyan Tang · Teresa Ruiz · Keith P Mintz ·
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    ABSTRACT: Aggregatibacter actinomycetemcomitans is hypothesized to colonize through the interaction with collagen and establish a reservoir for further dissemination. The trimeric adhesin EmaA of A. actinomycetemcomitans binds to collagen and is modified with sugars mediated by an O-antigen polysaccharide ligase (WaaL) that is associated with lipopolysaccharide (LPS) biosynthesis (G. Tang and K. Mintz, J. Bacteriol. 192:1395-1404, 2010). This investigation characterized the function and cellular localization of EmaA glycosylation. The interruption of LPS biogenesis by using genetic and pharmacological methods changed the amount and biophysical properties of EmaA molecules in the outer membrane. In rmlC and waaL mutant strains, the membrane-associated EmaA was reduced by 50% compared with the wild-type strain, without changes in mRNA levels. The membrane-associated EmaA protein levels were recovered by complementation with the corresponding O-polysaccharide (O-PS) biosynthetic genes. In contrast, another trimeric autotransporter, epithelial adhesin ApiA, was not affected in the same mutant background. The inhibition of undecaprenyl pyrophosphate recycling by bacitracin resulted in a similar decrease in the membrane-associated EmaA protein. This effect was reversed by removal of the compound. A significant decrease in collagen binding activity was observed in strains expressing the nonglycosylated form of EmaA. Furthermore, the electrophoretic mobility shifts of the EmaA monomers found in the O-PS mutant strains were associated only with the membrane-associated protein and not with the cytoplasmic pre-EmaA protein, suggesting that this modification does not occur in the cytoplasm. The glycan modification of EmaA appears to be required for collagen binding activity and protection of the protein against degradation by proteolytic enzymes.
    Infection and immunity 06/2012; 80(8):2868-77. DOI:10.1128/IAI.00372-12 · 3.73 Impact Factor
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    ABSTRACT: Adhesion to collagen is an important virulence determinant for the periodontal pathogen Aggregatibacter actinomycetemcomitans. Binding to collagen is mediated by the extracellular-matrix protein adhesin-A (EmaA). EmaA is a homotrimeric autotransporter protein that forms flexible antenna-like appendages on the bacterium surface. An ellipsoidal structure at the distal end of the appendage, composed of three subdomains, contains the functional domain of the molecule. A correlation between amino-acid sequence and subdomain structure (SI and SII) was proposed based on an analysis of the volume/molecular weight ratio. EmaA from three mutant strains (deletions of amino-acids 70-206 and 70-386 and a substitution mutation G162S) has been studied by electron microscopy to test this hypothesis. 3D structures were analyzed using single-axis tilt tomography of negatively stained preparations of bacteria combined with subvolume averaging. Additionally, a large number of 2D images of the apical domain of the adhesins from the mutants were extracted from micrographs of the bacterial surface, aligned and classified. The combined data showed that amino-acids 70-206 localize to subdomain SI and 70-386 comprise subdomains SI and SII. Moreover, we showed that the substitution mutation G162S, which abolishes collagen binding activity, does not affect the overall structural integrity of the functional domain. However, the structure of subdomain SI in this mutant is slightly altered with respect to the wild-type strain. These data also have allowed us to interpret the architectural features of each subdomain of EmaA in more detail and to correlate the 3D structure of the functional domain of EmaA with the amino-acid sequence.
    Journal of Structural Biology 12/2011; 177(2):439-46. DOI:10.1016/j.jsb.2011.11.024 · 3.23 Impact Factor
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    X Jiang · T Ruiz · K P Mintz ·
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    ABSTRACT: The extracellular matrix protein adhesin A (EmaA) of the Gram-negative bacterium Aggregatibacter actinomycetemcomitans is a fibrillar collagen adhesin belonging to the family of trimeric autotransporters. The protein forms antenna-like structures on the bacterial surface required for collagen adhesion. The 202-kDa protein monomers are proposed to be targeted and translocated across the inner membrane by a long signal peptide composed of 56 amino acids. The predicted signal peptide was functionally active in Escherichia coli and A. actinomycetemcomitans using truncated PhoA and Aae chimeric proteins, respectively. Mutations in the signal peptide were generated and characterized for PhoA activity in E. coli. A. actinomycetemcomitans strains expressing EmaA with the identical mutant signal peptides were assessed for cellular localization, surface expression, and collagen binding activity. All of the mutants impaired some aspect of EmaA structure or function. A signal peptide mutant that promoted alkaline phosphatase secretion did not allow any cell surface presentation of EmaA. A second mutant allowed for cell surface exposure but abolished protein function. A third mutant allowed for the normal localization and function of EmaA at 37°C but impaired localization at elevated temperatures. Likewise, replacement of the long EmaA signal peptide with a typical signal peptide also impaired localization above 37°C. The data suggest that the residues of the EmaA signal peptide are required for protein folding or assembly of this collagen adhesin.
    Journal of bacteriology 12/2011; 193(24):6983-94. DOI:10.1128/JB.05813-11 · 2.81 Impact Factor
  • S. Benjamin · M. Radermacher · T. Ruiz ·

    Microscopy and Microanalysis 07/2011; 17:80-81. DOI:10.1017/S1431927611001279 · 1.88 Impact Factor
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    Microscopy and Microanalysis 07/2011; 17(S2):90-91. DOI:10.1017/S1431927611001334 · 1.88 Impact Factor
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    Xiaobo Liang · Yi-Ywan M Chen · Teresa Ruiz · Hui Wu ·
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    ABSTRACT: Dental biofilm formation is critical for maintaining the healthy microbial ecology of the oral cavity. Streptococci are predominant bacterial species in the oral cavity and play important roles in the initiation of plaque formation. In this study, we identified a new cell surface protein, BapA1, from Streptococcus parasanguinis FW213 and determined that BapA1 is critical for biofilm formation. Sequence analysis revealed that BapA1 possesses a typical cell wall-sorting signal for cell surface-anchored proteins from Gram-positive bacteria. No functional orthologue was reported in other streptococci. BapA1 possesses nine putative pilin isopeptide linker domains which are crucial for pilus assembly in a number of Gram-positive bacteria. Deletion of the 3' portion of bapA1 generated a mutant that lacks surface-anchored BapA1 and abolishes formation of short fibrils on the cell surface. The mutant failed to form biofilms and exhibited reduced adherence to an in vitro tooth model. The BapA1 deficiency also inhibited bacterial autoaggregation. The N-terminal muramidase-released-protein-like domain mediated BapA1-BapA1 interactions, suggesting that BapA1-mediated cell-cell interactions are important for bacterial autoaggregation and biofilm formation. Furthermore, the BapA1-mediated bacterial adhesion and biofilm formation are independent of a fimbria-associated serine-rich repeat adhesin, Fap1, demonstrating that BapA1 is a new streptococcal adhesin.
    Infection and immunity 05/2011; 79(8):3239-48. DOI:10.1128/IAI.00029-11 · 3.73 Impact Factor

Publication Stats

1k Citations
179.87 Total Impact Points


  • 2004-2015
    • University of Vermont
      • • Department of Molecular Physiology and Biophysics
      • • Department of Microbiology and Molecular Genetics
      • • College of Medicine
      Burlington, Vermont, United States
  • 2003
    • University of Vermont Medical Center
      Burlington, Vermont, United States
  • 2002
    • Goethe-Universität Frankfurt am Main
      Frankfurt, Hesse, Germany
  • 2001-2002
    • Max Planck Institute of Biophysics
      • Department of Molecular Membrane Biology
      Frankfurt, Hesse, Germany