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Plasmid-encoded outer membrane protein YadA mediates specific binding of enteropathogenic yersiniae to various types
H Schulze-Koops1, H Burkhardt, J Heesemann
1Max-Planck-Society, University Erlangen-Nürnberg, Germany.
Abstract:
The plasmid-encoded outer membrane protein YadA of enteropathogenic yersiniae is associated with pathogenicity. Recently, collagen binding of YadA-positive yersiniae was reported without detailed characterization (L. Emödy, J. Heesemann, H. Wolf-Watz, M. Skurnik, G. Kapperud, P. O'Toole, and T. Wadström, J. Bacteriol. 171:6674-6679, 1989). To elucidate the nature of collagen binding to YadA, we used a recombinant Yersinia strain expressing the cloned YadA gene. Direct binding of YadA-positive yersiniae to collagens was demonstrated in affinity blot experiments on nitrocellulose filters. A spectrum of collagen types in a wide concentration range were tested for their ability to block binding of 125I-labeled collagen type II to YadA-positive yersiniae. The results indicate a specific binding site(s) for YadA in collagen types I, II, III, IV, V, and XI. In contrast, collagen type VI did not bind to YadA. To characterize the binding site(s) more precisely, isolated collagen chains and cyanogen bromide fragments were investigated. These studies revealed that binding of YadA to collagen type I is confined to the alpha 1(I) chain, whereas the binding site within collagen type XI is localized in the alpha 3(XI) chain. alpha 2(I), alpha 1(XI), and alpha 2(XI) did not bind to YadA. Most interestingly, in the alpha 1(II) chain the specific binding site for YadA resides in the cyanogen bromide fragment CB10. The latter might indicate a binding site that does not depend on conformation. Based on these findings, further fragmentation and the synthesis of peptides may allow definition of the peptide sequence(s) relevant for YadA binding.
Insights
The outer membrane protein YadA from enteropathogenic yersiniae specifically binds to multiple collagen types. Researchers identified specific binding sites within collagen chains, paving the way for defining YadA-collagen interaction sequences.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- The plasmid-encoded outer membrane protein YadA is linked to the pathogenicity of enteropathogenic yersiniae.
- Previous reports noted collagen binding by YadA-positive yersiniae without detailed characterization.
Purpose of the Study:
- To elucidate the specific nature of collagen binding to the YadA protein.
- To identify the collagen types and specific chains/fragments that interact with YadA.
Main Methods:
- Utilized a recombinant Yersinia strain expressing the cloned YadA gene.
- Performed direct binding assays using affinity blot experiments on nitrocellulose filters.
- Investigated binding to various collagen types, isolated collagen chains, and cyanogen bromide fragments.
Main Results:
- Demonstrated direct binding of YadA-positive yersiniae to collagens I, II, III, IV, V, and XI, but not type VI.
- Identified specific binding of YadA to the alpha 1(I) chain of collagen type I and the alpha 3(XI) chain of collagen type XI.
- Localized the YadA binding site on the alpha 1(II) chain of collagen type II to the cyanogen bromide fragment CB10.
Conclusions:
- YadA exhibits specific binding interactions with multiple collagen types.
- The binding site for YadA on collagen type II (fragment CB10) may be conformation-independent.
- Further studies involving peptide synthesis can define the precise peptide sequences involved in YadA-collagen binding.