Prevalence of ompT among Escherichia coli isolates of human origin

Lundrigan, Michael D.; Webb, Richard A.

doi:10.1016/0378-1097(92)90362-r

Cited by 32 publications

(23 citation statements)

References 19 publications

(15 reference statements)

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“…OmpT is a serine protease well characterized to be an important virulence factor in NMEC and UPEC strains (78)(79)(80)(81)(82). In STEC, OmpT degrades LL-37 (83), an antimicrobial peptide secreted by epithelial cells of the stomach and colon (84,85).…”

Section: Discussionmentioning

confidence: 99%

Immunoproteomic Analysis To Identify Shiga Toxin-Producing Escherichia coli Outer Membrane Proteins Expressed during Human Infection

et al. 2014

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b Shiga-toxin producing Escherichia coli (STEC) is the etiologic agent of acute diarrhea, dysentery, and hemolytic-uremic syndrome (HUS). There is no approved vaccine for STEC infection in humans, and antibiotic use is contraindicated, as it promotes Shiga toxin production. In order to identify STEC-associated antigens and immunogenic proteins, outer membrane proteins (OMPs) were extracted from STEC O26:H11, O103, O113:H21, and O157:H7 strains, and commensal E. coli strain HS was used as a control. SDS-PAGE, two-dimensional-PAGE analysis, Western blot assays using sera from pediatric HUS patients and controls, and matrix-assisted laser desorption ionization-tandem time of flight analyses were used to identify 12 immunogenic OMPs, some of which were not reactive with control sera. Importantly, seven of these proteins have not been previously reported to be immunogenic in STEC strains. Among these seven proteins, OmpT and Cah displayed IgG and IgA reactivity with sera from HUS patients. Genes encoding these two proteins were present in a majority of STEC strains. Knowledge of the antigens produced during infection of the host and the immune response to those antigens will be important for future vaccine development.

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Section: Discussionmentioning

confidence: 99%

Immunoproteomic Analysis To Identify Shiga Toxin-Producing Escherichia coli Outer Membrane Proteins Expressed during Human Infection

et al. 2014

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“…More than 20 years ago, Leytus and coworkers showed that OmpT catalyzes the activation of plasminogen to plasmin (17), a function that is physiologically relevant for the virulence of Yersinia pestis and for clinical E. coli isolates (18,28). OmpT has also been found to play a role in bacterial virulence in ways that are unrelated to plasminogen activation, for example in the cleavage of protamine and other cationic peptides with antibiotic activity (11,29).…”

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confidence: 99%

Substrate Specificity of the Escherichia coli Outer Membrane Protease OmpT

McCarter

Stephens

Shoemaker³

et al. 2004

J Bacteriol

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OmpT is a surface protease of gram-negative bacteria that has been shown to cleave antimicrobial peptides, activate human plasminogen, and degrade some recombinant heterologous proteins. We have analyzed the substrate specificity of OmpT by two complementary substrate filamentous phage display methods: (i) in situ cleavage of phage that display protease-susceptible peptides by Escherichia coli expressing OmpT and (ii) in vitro cleavage of phage-displayed peptides using purified enzyme. Consistent with previous reports, OmpT was found to exhibit a virtual requirement for Arg in the P1 position and a slightly less stringent preference for this residue in the P1 position (P1 and P1 are the residues immediately prior to and following the scissile bond). Lys, Gly, and Val were also found in the P1 position. The most common residues in the P2 position were Val or Ala, and the P3 and P4 positions exhibited a preference for Trp or Arg. Synthetic peptides based upon sequences selected by bacteriophage display were cleaved very efficiently, with k cat /K m values up to 7.3 ؋ 10 6 M ؊1 s ؊1 . In contrast, a peptide corresponding to the cleavage site of human plasminogen was hydrolyzed with a k cat /K m almost 10 6 -fold lower. Overall, the results presented in this work indicate that in addition to the P1 and P1 positions, additional amino acids within a six-residue window (between P4 and P2) contribute to the binding of substrate polypeptides to the OmpT binding site.

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“…5B). However, the wildtype enzyme was not specific for the monobasic cleavage site on PAC and generated not only ACTH but also the by-products RAR-ACTH, ACTH (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14)(15), and ACTH (16)(17)(18)(19)(20)(21)(22)(23)(24). On the other hand, the OmpT variant D97L released ACTH (1-24) specifically and efficiently, which coincides well with its substrate specificity for the fusion protein PRS instead of PRR ( Table 1).…”

Section: Discussionmentioning

confidence: 99%

“…Wild-type OmpT also released ACTH , but the by-product RAR-ACTH was generated from cleavage between Arg 140 and Arg 141 . Additionally, ACTH (1-15) and ACTH (16)(17)(18)(19)(20)(21)(22)(23)(24) were generated by cleavage at Arg 143 -Ser 144 and Lys 158 -Lys 159 (data not shown). When the variant OmpT and wild-type OmpT were compared, the amount of ACTH (1-24) released by OmpT variant D97L was 2.9-fold higher than the amount released by the wild-type OmpT.…”

Section: Vol 70 2004 Utilization Of Ompt Variants As Processing Enzmentioning

confidence: 99%

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Utilization of Escherichia coli Outer-Membrane Endoprotease OmpT Variants as Processing Enzymes for Production of Peptides from Designer Fusion Proteins

Okuno

Yabuta

Ooi

et al. 2004

Appl Environ Microbiol

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Escherichia coli outer-membrane endoprotease OmpT has suitable properties for processing fusion proteins to produce peptides and proteins. However, utilization of this protease for such production has been restricted due to its generally low cleavage efficiency at Arg (or Lys)-Xaa, where Xaa is a nonbasic N-terminal amino acid of a target polypeptide. The objective of this study was to generate a specific and efficient OmpT protease and to utilize it as a processing enzyme for producing various peptides and proteins by converting its substrate specificity. Since OmpT Asp 97 is proposed to interact with the P1 amino acid of its substrates, OmpT variants with variations at Asp 97 were constructed by replacing this amino acid with 19 natural amino acids to alter the cleavage specificity at Arg (P1)-Xaa (P1). The variant OmpT that had a methionine at this position, but not the wild-type OmpT, efficiently cleaved a fusion protein containing the amino acid sequence -Arg-Arg-ArgAla-Arg2motilin, in which motilin is a model peptide with a phenylalanine at the N terminus. The OmpT variants with leucine and histidine at position 97 were useful in releasing human adrenocorticotropic hormone (1-24) (serine at the N terminus) and human calcitonin precursor (cysteine at the N terminus), respectively, from fusion proteins. Motilin was produced by this method and was purified up to 99.0% by two chromatographic steps; the yield was 160 mg/liter of culture. Our novel method in which the OmpT variants are used could be employed for production of various peptides and proteins.

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Prevalence of ompT among Escherichia coli isolates of human origin

Cited by 32 publications

References 19 publications

Immunoproteomic Analysis To Identify Shiga Toxin-Producing Escherichia coli Outer Membrane Proteins Expressed during Human Infection

Immunoproteomic Analysis To Identify Shiga Toxin-Producing Escherichia coli Outer Membrane Proteins Expressed during Human Infection

Substrate Specificity of the Escherichia coli Outer Membrane Protease OmpT

Utilization of Escherichia coli Outer-Membrane Endoprotease OmpT Variants as Processing Enzymes for Production of Peptides from Designer Fusion Proteins

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