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Rational Design and Evaluation of an Artificial Escherichia coli K1 Protein Vaccine Candidate Based on the Structure of OmpA.

ABSTRACT: Escherichia coli (E. coli) K1 causes meningitis and remains an unsolved problem in neonates, despite the application of antibiotics and supportive care. The cross-reactivity of bacterial capsular polysaccharides with human antigens hinders their application as vaccine candidates. Thus, protein antigens could be an alternative strategy for the development of an E. coli K1 vaccine. Outer membrane protein A (OmpA) of E. coli K1 is a potential vaccine candidate because of its predominant contribution to bacterial pathogenesis and sub-cellular localization. However, little progress has been made regarding the use of OmpA for this purpose due to difficulties in OmpA production. In the present study, we first investigated the immunogenicity of the four extracellular loops of OmpA. Using the structure of OmpA, we rationally designed and successfully generated the artificial protein OmpAVac, composed of connected loops from OmpA. Recombinant OmpAVac was successfully produced in E. coli BL21 and behaved as a soluble homogenous monomer in the aqueous phase. Vaccination with OmpAVac induced Th1, Th2, and Th17 immune responses and conferred effective protection in mice. In addition, OmpAVac-specific antibodies were able to mediate opsonophagocytosis and inhibit bacterial invasion, thereby conferring prophylactic protection in E. coli K1-challenged adult mice and neonatal mice. These results suggest that OmpAVac could be a good vaccine candidate for the control of E. coli K1 infection and provide an additional example of structure-based vaccine design.

SUBMITTER: Gu H

PROVIDER: S-EPMC5974202 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Rational Design and Evaluation of an Artificial Escherichia coli K1 Protein Vaccine Candidate Based on the Structure of OmpA.

Gu Hao H Liao Yaling Y Zhang Jin J Wang Ying Y Liu Zhiyong Z Cheng Ping P Wang Xingyong X Zou Quanming Q Gu Jiang J

Frontiers in cellular and infection microbiology 20180523

Escherichia coli (E. coli) K1 causes meningitis and remains an unsolved problem in neonates, despite the application of antibiotics and supportive care. The cross-reactivity of bacterial capsular polysaccharides with human antigens hinders their application as vaccine candidates. Thus, protein antigens could be an alternative strategy for the development of an E. coli K1 vaccine. Outer membrane protein A (OmpA) of E. coli K1 is a potential vaccine candidate because of ...[more]

PMID: 29876324

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Project description:E. coli O157:H7, one of the major EHEC serotypes, is capable of developing bloody diarrhea, hemorrhagic colitis (HC), and fatal hemolytic uremic syndrome (HUS) and is accompanied by high annual economic loss worldwide. Due to the increased risk of HC and HUS development following antibiotic therapy, the prevention of infections caused by this pathogen is considered to be one of the most effective ways of avoiding the consequences of this infection. The main aim of the present study was to design, express, and purify a novel chimeric protein to develope human vaccine candidate against E. coli O157:H7 containing loop 2-4 of E. coli O157:H7, outer membrane protein A (OmpA), and B subunit of E. coli heat labile enterotoxin (LTB) which are connected by a flexible peptide linker. Several online databases and bioinformatics software were utilized to choose the peptide linker among 537 analyzed linkers, design the chimeric protein, and optimize the codon of the relative gene encoding this protein. Subsequently, the recombinant gene encoding OmpA-LTB was synthesized and cloned into pET-24a (+) expression vector and transferred to E. coli BL21(DE3) cells. The expression of OmpA-LTB chimeric protein was then carried out by induction of cultured E. coli Bl21 (DE3) cells with 1mM isopropyl-β-D-thiogalactopyranoside (IPTG). The purification of OmpA-LTB was then performed by nickel affinity chromatography. Expression and purification were analyzed by sodium dodecyl sulphate poly acrylamide gel electrophoresis. Moreover, the identity of the expressed protein was analyzed by western blotting. SDS-PAGE and western immunoblotting confirmed the successful expression of a 27 KDa recombinant protein after 24 hours at 37°C post-IPTG induction. OmpA-LTB was then successfully purified, using nickel affinity chromatography under denaturing conditions. The yield of purification was 12 mg per liter of culture media. Ultimately, we constructed the successful design and efficient expression and purification of OmpA-LTB divalent under the above-mentioned conditions.

Dataset Information

Rational Design and Evaluation of an Artificial Escherichia coli K1 Protein Vaccine Candidate Based on the Structure of OmpA.

Publications

Rational Design and Evaluation of an Artificial <i>Escherichia coli</i> K1 Protein Vaccine Candidate Based on the Structure of OmpA.

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