scholarly journals Periplasmic Peptidyl Prolyl cis-trans Isomerases Are Not Essential for Viability, but SurA Is Required for Pilus Biogenesis in Escherichia coli

2005 ◽  
Vol 187 (22) ◽  
pp. 7680-7686 ◽  
Author(s):  
Sheryl S. Justice ◽  
David A. Hunstad ◽  
Jill Reiss Harper ◽  
Amy R. Duguay ◽  
Jerome S. Pinkner ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane Proteins ◽  
E Coli ◽  
Outer Membranes ◽  
Type 1 Pili ◽  
Pilus Biogenesis ◽  
Quadruple Mutant ◽  
Increased Susceptibility ◽  
Pilus Assembly

ABSTRACT In Escherichia coli, FkpA, PpiA, PpiD, and SurA are the four known periplasmic cis-trans prolyl isomerases. These isomerases facilitate proper protein folding by increasing the rate of transition of proline residues between the cis and trans states. Genetic inactivation of all four periplasmic isomerases resulted in a viable strain that exhibited a decreased growth rate and increased susceptibility to certain antibiotics. Levels of the outer membrane proteins LamB and OmpA in the quadruple mutant were indistinguishable from those in the surA single mutant. In addition, expression of P and type 1 pili (adhesive organelles produced by uropathogenic strains of E. coli and assembled by the chaperone/usher pathway) were severely diminished in the absence of the four periplasmic isomerases. Maturation of the usher was significantly impaired in the outer membranes of strains devoid of all four periplasmic isomerases, resulting in a defect in pilus assembly. Moreover, this defect in pilus assembly and usher stability could be attributed to the absence of SurA. The data presented here suggest that the four periplasmic isomerases are not essential for growth under laboratory conditions but may have significant roles in survival in environmental and pathogenic niches, as indicated by the effect on pilus production.

scholarly journals Chaperone-Subunit-Usher Interactions Required for Donor Strand Exchange during Bacterial Pilus Assembly

2003 ◽  
Vol 185 (9) ◽  
pp. 2723-2730 ◽  
Author(s):  
Michelle M. Barnhart ◽  
Frederic G. Sauer ◽  
Jerome S. Pinkner ◽  
Scott J. Hultgren
Keyword(s):  
Escherichia Coli ◽  
Outer Membrane ◽  
Strand Exchange ◽  
Assembly Mechanism ◽  
Type 1 Pili ◽  
Pilus Biogenesis ◽  
A Subunit ◽  
Donor Strand Exchange ◽  
Pilus Assembly

ABSTRACT The assembly of type 1 pili on the surface of uropathogenic Escherichia coli proceeds via the chaperone-usher pathway. Chaperone-subunit complexes interact with one another via a process termed donor strand complementation whereby the G1β strand of the chaperone completes the immunoglobulin (Ig) fold of the pilus subunit. Chaperone-subunit complexes are targeted to the usher, which forms a channel across the outer membrane through which pilus subunits are translocated and assembled into pili via a mechanism known as donor strand exchange. This is a mechanism whereby chaperone uncapping from a subunit is coupled with the simultaneous assembly of the subunit into the pilus fiber. Thus, in the pilus fiber, the N-terminal extension of every subunit completes the Ig fold of its neighboring subunit by occupying the same site previously occupied by the chaperone. Here, we investigated details of the donor strand exchange assembly mechanism. We discovered that the information necessary for targeting the FimC-FimH complex to the usher resides mainly in the FimH protein. This interaction is an initiating event in pilus biogenesis. We discovered that the ability of an incoming subunit (in a chaperone-subunit complex) to participate in donor strand exchange with the growing pilus depended on a previously unrecognized function of the chaperone. Furthermore, the donor strand exchange assembly mechanism between subunits was found to be necessary for subunit translocation across the outer membrane usher.

scholarly journals Processive dynamics of the usher assembly platform during uropathogenic Escherichia coli P pilus biogenesis

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Minge Du ◽  
Zuanning Yuan ◽  
Glenn T. Werneburg ◽  
Nadine S. Henderson ◽  
Hemil Chauhan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Bacterial Colonization ◽  
Conformational Dynamics ◽  
Structural Heterogeneity ◽  
Structural Basis ◽  
Type 1 Pili ◽  
Pilus Biogenesis ◽  
Functional Understanding ◽  
Pilus Assembly

AbstractUropathogenic Escherichia coli assemble surface structures termed pili or fimbriae to initiate infection of the urinary tract. P pili facilitate bacterial colonization of the kidney and pyelonephritis. P pili are assembled through the conserved chaperone-usher pathway. Much of the structural and functional understanding of the chaperone-usher pathway has been gained through investigations of type 1 pili, which promote binding to the bladder and cystitis. In contrast, the structural basis for P pilus biogenesis at the usher has remained elusive. This is in part due to the flexible and variable-length P pilus tip fiber, creating structural heterogeneity, and difficulties isolating stable P pilus assembly intermediates. Here, we circumvent these hindrances and determine cryo-electron microscopy structures of the activated PapC usher in the process of secreting two- and three-subunit P pilus assembly intermediates, revealing processive steps in P pilus biogenesis and capturing new conformational dynamics of the usher assembly machine.

scholarly journals Processive Dynamics of the Usher Assembly Platform During Uropathogenic Escherichia coli P Pilus Biogenesis

2020 ◽  
Author(s):  
Minge Du ◽  
Zuanning Yuan ◽  
Glenn T. Werneburg ◽  
Nadine S. Henderson ◽  
Hemil Chauhan ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Bacterial Colonization ◽  
Conformational Dynamics ◽  
Structural Heterogeneity ◽  
Structural Basis ◽  
Bladder Epithelium ◽  
Type 1 Pili ◽  
Pilus Biogenesis ◽  
Pilus Assembly

ABSTRACTUropathogenic Escherichia coli (UPEC) assemble hair-like surface structures termed pili or fimbriae to initiate infection of the urinary tract. P pili mediate the adherence of UPEC to the kidney epithelium, facilitating bacterial colonization and pyelonephritis1. P pili are assembled through the conserved chaperone-usher (CU) pathway2-4. In this pathway, a dedicated chaperone facilitates the folding of nascent pilus subunits in the periplasm and an integral outer membrane (OM) protein termed the usher provides the assembly platform and secretion channel for the pilus fiber. Much of the structural and functional understanding of the CU pathway has been gained through investigations of type 1 pili, which promote UPEC binding to the bladder epithelium and the development of cystitis5. In contrast, the structural basis for P pilus biogenesis at the usher has remained elusive. This is in part due to the flexible and variable-length P pilus tip fiber, creating structural heterogeneity, as well as difficulties in isolating stable P pilus assembly intermediates from bacteria. Here, we have devised a method to circumvent these hindrances and determined cryo-EM structures of the activated PapC usher in the process of secreting two- and three-subunit P pilus assembly intermediates. These structures show processive steps in P pilus biogenesis, reveal differences between P and type 1 pili, and capture new conformational dynamics of the usher assembly machine.

scholarly journals The Escherichia coli P and Type 1 Pilus Assembly Chaperones PapD and FimC Are Monomeric in Solution

2016 ◽  
Vol 198 (17) ◽  
pp. 2360-2369 ◽  
Author(s):  
Samema Sarowar ◽  
Olivia J. Hu ◽  
Glenn T. Werneburg ◽  
David G. Thanassi ◽  
Huilin Li
Keyword(s):  
Escherichia Coli ◽  
Oligomeric State ◽  
Content Type ◽  
Dimer Interface ◽  
Adhesive Surface ◽  
Type 1 Pili ◽  
Pilus Biogenesis ◽  
Multistep Process ◽  
Pilus Assembly

ABSTRACTThe chaperone/usher pathway is used by Gram-negative bacteria to assemble adhesive surface structures known as pili or fimbriae. Uropathogenic strains ofEscherichia coliuse this pathway to assemble P and type 1 pili, which facilitate colonization of the kidney and bladder, respectively. Pilus assembly requires a periplasmic chaperone and outer membrane protein termed the usher. The chaperone allows folding of pilus subunits and escorts the subunits to the usher for polymerization into pili and secretion to the cell surface. Based on previous structures of mutant versions of the P pilus chaperone PapD, it was suggested that the chaperone dimerizes in the periplasm as a self-capping mechanism. Such dimerization is counterintuitive because the chaperone G1 strand, important for chaperone-subunit interaction, is buried at the dimer interface. Here, we show that the wild-type PapD chaperone also forms a dimer in the crystal lattice; however, the dimer interface is different from the previously solved structures. In contrast to the crystal structures, we found that both PapD and the type 1 pilus chaperone, FimC, are monomeric in solution. Our findings indicate that pilus chaperones do not sequester their G1 β-strand by forming a dimer. Instead, the chaperones may expose their G1 strand for facile interaction with pilus subunits. We also found that the type 1 pilus adhesin, FimH, is flexible in solution while in complex with its chaperone, whereas the P pilus adhesin, PapGII, is rigid. Our study clarifies a crucial step in pilus biogenesis and reveals pilus-specific differences that may relate to biological function.IMPORTANCEPili are critical virulence factors for many bacterial pathogens. UropathogenicE. colirelies on P and type 1 pili assembled by the chaperone/usher pathway to adhere to the urinary tract and establish infection. Studying pilus assembly is important for understanding mechanisms of protein secretion, as well as for identifying points for therapeutic intervention. Pilus biogenesis is a multistep process. This work investigates the oligomeric state of the pilus chaperone in the periplasm, which is important for understanding early assembly events. Our work unambiguously demonstrates that both PapD and FimC chaperones are monomeric in solution. We further demonstrate that the solution behavior of the FimH and PapGII adhesins differ, which may be related to functional differences between the two pilus systems.

scholarly journals Immunoglobulin-Mediated Agglutination of and Biofilm Formation by Escherichia coli K-12 Require the Type 1 Pilus Fiber

2004 ◽  
Vol 72 (4) ◽  
pp. 1929-1938 ◽  
Author(s):  
Paul E. Orndorff ◽  
Aditya Devapali ◽  
Sarah Palestrant ◽  
Aaron Wyse ◽  
Mary Lou Everett ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Biofilm Formation ◽  
Immunoglobulin A ◽  
Minor Component ◽  
Secretory Iga ◽  
Bacterial Surface ◽  
E Coli ◽  
Type 1 Pili ◽  
K 12

ABSTRACT The binding of human secretory immunoglobulin A (SIgA), the primary immunoglobulin in the gut, to Escherichia coli is thought to be dependent on type 1 pili. Type 1 pili are filamentous bacterial surface attachment organelles comprised principally of a single protein, the product of the fimA gene. A minor component of the pilus fiber (the product of the fimH gene, termed the adhesin) mediates attachment to a variety of host cell molecules in a mannose inhibitable interaction that has been extensively described. We found that the aggregation of E. coli K-12 by human secretory IgA (SIgA) was dependent on the presence of the pilus fiber, even in the absence of the mannose specific adhesin or in the presence of 25 mM α-CH3Man. The presence of pilus without adhesin also facilitated SIgA-mediated biofilm formation on polystyrene, although biofilm formation was stronger in the presence of the adhesin. IgM also mediated aggregation and biofilm formation in a manner dependent on pili with or without adhesin. These findings indicate that the pilus fiber, even in the absence of the adhesin, may play a role in biologically important processes. Under conditions in which E. coli was agglutinated by SIgA, the binding of SIgA to E. coli was not increased by the presence of the pili, with or without adhesin. This observation suggests that the pili, with or without adhesin, affect factors such as cell surface rigidity or electrostatic repulsion, which can affect agglutination but which do not necessarily determine the level of bound immunoglobulin.

scholarly journals A Novel Putrescine Importer Required for Type 1 Pili-driven Surface Motility Induced by Extracellular Putrescine in Escherichia coli K-12

2011 ◽  
Vol 286 (12) ◽  
pp. 10185-10192 ◽  
Author(s):  
Shin Kurihara ◽  
Hideyuki Suzuki ◽  
Mayu Oshida ◽  
Yoshimi Benno
Keyword(s):  
Escherichia Coli ◽  
Plasmid Vector ◽  
The Other ◽  
Gene Deletions ◽  
E Coli ◽  
Type 1 Pili ◽  
Transport Assay ◽  
Surface Motility ◽  
K 12

Recently, many studies have reported that polyamines play a role in bacterial cell-to-cell signaling processes. The present study describes a novel putrescine importer required for induction of type 1 pili-driven surface motility. The surface motility of the Escherichia coli ΔspeAB ΔspeC ΔpotABCD strain, which cannot produce putrescine and cannot import spermidine from the medium, was induced by extracellular putrescine. Introduction of the gene deletions for known polyamine importers (ΔpotE, ΔpotFGHI, and ΔpuuP) or a putative polyamine importer (ΔydcSTUV) into the ΔspeAB ΔspeC ΔpotABCD strain did not affect putrescine-induced surface motility. The deletion of yeeF, an annotated putative putrescine importer, in the ΔspeAB ΔspeC ΔpotABCD ΔydcSTUV strain abolished surface motility in putrescine-supplemented medium. Complementation of yeeF by a plasmid vector restored surface motility. The surface motility observed in the present study was abolished by the deletion of fimA, suggesting that the surface motility is type 1 pili-driven. A transport assay using the yeeF+ or ΔyeeF strains revealed that YeeF is a novel putrescine importer. The Km of YeeF (155 μm) is 40 to 300 times higher than that of other importers reported previously. On the other hand, the Vmax of YeeF (9.3 nmol/min/mg) is comparable to that of PotABCD, PotFGHI, and PuuP. The low affinity of YeeF for putrescine may allow E. coli to sense the cell density depending on the concentration of extracellular putrescine.

scholarly journals Prevalence of Quinolone Resistance of Extended-Spectrum β-Lactamase-Producing Escherichia coli with ST131-fimH30 in a City Hospital in Hyogo, Japan

2019 ◽  
Vol 20 (20) ◽  
pp. 5162 ◽  
Author(s):  
Masazumi Teramae ◽  
Kayo Osawa ◽  
Katsumi Shigemura ◽  
Koichi Kitagawa ◽  
Toshiro Shirakawa ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Antibacterial Agents ◽  
City Hospital ◽  
Antimicrobial Susceptibility Test ◽  
E Coli ◽  
Specific Pcr ◽  
Extended Spectrum ◽  
Type 1 Pili ◽  
Genes Encoding

Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli isolates are known to tolerate superior quinolone antimicrobials compared with other antibacterial agents. Among the clones belonging to sequence type (ST) 131 by multilocus sequence typing, the involvement of the H30-Rx subclone has been reported worldwide with various fimH genes encoding type 1 pili. We investigated 83 isolates of ESBL-producing E. coli and performed antimicrobial susceptibility test, CH (fumC/fimH) ST131 by typing the specific PCR. Moreover, mutation analysis of genes involved in quinolone antibiotic resistance (gyrA and parC) and ESBL genotypes were determined. As a result, 54 of 83 isolates (65.1%) of CH40-30 clones corresponding to ST131-fimH30 were detected, and all were resistant to levofloxacin. Mutations associated with this resistance were common, and included S83L and D87N of gyrA and S80I and E84V of parC. Subclone analysis revealed a high proportion of fimH30-non-Rx (40 isolates, 74.1%). Each subclone was characterized by ESBL genotype, and the CTX-M-15 type was mainly seen for fimH30-Rx, with the CTX-M-14 type or CTX-M-27 type seen for fimH30-non-Rx. This study suggests that an increase in ESBL-producing quinolone-resistant E. coli in a city hospital in Hyogo, Japan, was caused by the spread of subclones belonging to fimH30-non-Rx of ST131.

scholarly journals Fimbrial Adhesins Produced by Atypical Enteropathogenic Escherichia coli Strains

2011 ◽  
Vol 77 (23) ◽  
pp. 8391-8399 ◽  
Author(s):  
Rodrigo T. Hernandes ◽  
Irina Velsko ◽  
Suely C. F. Sampaio ◽  
Waldir P. Elias ◽  
Roy M. Robins-Browne ◽  
...  
Keyword(s):  
Escherichia Coli ◽  
Phenotypic Expression ◽  
Complex Nature ◽  
Content Type ◽  
E Coli ◽  
Type 1 Pili ◽  
Gut Mucosa ◽  
Fimbrial Adhesins ◽  
Laminin Binding

ABSTRACTAtypical enteropathogenicEscherichia coli(aEPEC) has emerged as a significant cause of pediatric diarrhea worldwide; however, information regarding its adherence mechanisms to the human gut mucosa is lacking. In this study, we investigated the prevalence of several (fimA,ecpA,csgA,elfA, andhcpA) fimbrial genes in 71 aEPEC strains isolated from children with diarrhea (54 strains) and healthy individuals (17 strains) in Brazil and Australia by PCR. These genes are associated with adhesion and/or biofilm formation of pathogenic and commensalE. coli. Here, the most prevalent fimbrial genes found, in descending order, werehcpA(98.6%),ecpA(86%),fimA(76%),elfA(72%), andcsgA(19.7%). Phenotypic expression of pili in aEPEC strains was assessed by several approaches. We were not able to detect the hemorrhagic coli pilus (HCP) or theE. colilaminin-binding fimbriae (ELF) in these strains by using immunofluorescence. Type 1 pili and curli were detected in 59% (by yeast agglutination) and 2.8% (by Congo red binding and immunofluorescence) of the strains, respectively. TheE. colicommon pilus (ECP) was evidenced in 36.6% of the strains on bacteria adhering to HeLa cells by immunofluorescence, suggesting that ECP could play an important role in cell adherence for some aEPEC strains. This study highlights the complex nature of the adherence mechanisms of aEPEC strains involving the coordinated function of fimbrial (e.g., ECP) and nonfimbrial (e.g., intimin) adhesins and indicates that these strains bear several pilus operons that could potentially be expressed in different niches favoring colonization and survival in and outside the host.

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