Heterologous Expression of Bartonella Adhesin A in Escherichia coli by Exchange of Trimeric Autotransporter Adhesin Domains Results in Enhanced Adhesion Properties and a Pathogenic Phenotype

Author:

Schmidgen Thomas1,Kaiser Patrick O.1,Ballhorn Wibke1,Franz Bettina1,Göttig Stephan1,Linke Dirk23,Kempf Volkhard A. J.1

Affiliation:

1. Institut für Medizinische Mikrobiologie und Krankenhaushygiene, Klinikum der Goethe-Universität, Frankfurt am Main, Germany

2. Max-Planck-Institut für Entwicklungsbiologie, Abteilung Proteinevolution, Tübingen, Germany

3. Department of Biosciences, University of Oslo, Oslo, Norway

Abstract

ABSTRACT Human-pathogenic Bartonella henselae causes cat scratch disease and vasculoproliferative disorders. An important pathogenicity factor of B. henselae is the trimeric autotransporter adhesin (TAA) Bartonella adhesin A (BadA), which is modularly constructed, consisting of a head, a long and repetitive neck-stalk module, and a membrane anchor. BadA is involved in bacterial autoagglutination, binding to extracellular matrix proteins and host cells, and in proangiogenic reprogramming. The slow growth of B. henselae and limited tools for genetic manipulation are obstacles for detailed examination of BadA and its domains. Here, we established a recombinant expression system for BadA mutants in Escherichia coli allowing functional analysis of particular BadA domains. Using a BadA mutant lacking 21 neck-stalk repeats (BadA HN23), the BadA HN23 signal sequence was exchanged with that of E. coli OmpA, and the BadA membrane anchor was additionally replaced with that of Yersinia adhesin A (YadA). Constructs were cloned in E. coli , and hybrid protein expression was detected by immunoblotting, fluorescence microscopy, and flow cytometry. Functional analysis revealed that BadA hybrid proteins mediate autoagglutination and binding to collagen and endothelial cells. In vivo , expression of this BadA construct correlated with higher pathogenicity of E. coli in a Galleria mellonella infection model.

Publisher

American Society for Microbiology

Subject

Molecular Biology,Microbiology

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