Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex

Niemann H, Gherardi E, Bleymüller W, Heinz DW (2012)
Protein Science 21(10): 1528-1539.

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Abstract
The physiological relevance of contacts in crystal lattices often remains elusive. This was also the case for the complex between the invasion protein internalin B (InlB) from Listeria monocytogenes and its host cell receptor, the human receptor tyrosine kinase (RTK) MET. InlB is a MET agonist and induces bacterial host cell invasion. Activation of RTKs generally involves ligand-induced dimerization of the receptor ectodomain. The two currently available crystal structures of the InlB:MET complex show the same arrangement of InlB and MET in a 1:1 complex, but different dimeric 2:2 assemblies. Only one of these 2:2 assemblies is predicted to be stable by a computational procedure. This assembly is mainly stabilized by a contact between the Cap domain of InlB from one and the Sema domain of MET from another 1:1 complex. Here, we probe the physiological relevance of this interaction. We generated variants of the leucine-rich repeat (LRR) protein InlB by inserting an additional repeat between the first and the second LRR. This should allow formation of the 1:1 complex but disrupt the potential 2:2 complex involving the Cap-Sema contact due to steric distortions. A crystal structure of one of the engineered proteins showed that it folded properly. Binding affinity to MET was comparable to that of wild-type InlB. The InlB variant induced MET phosphorylation and cell scatter like wild-type InlB. These results suggest that the Cap-Sema interaction is not physiologically relevant and support the previously proposed assembly, in which a 2:2 InlB:MET complex is built around a ligand dimer. Copyright 2012 The Protein Society.
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Niemann H, Gherardi E, Bleymüller W, Heinz DW. Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex. Protein Science. 2012;21(10):1528-1539.
Niemann, H., Gherardi, E., Bleymüller, W., & Heinz, D. W. (2012). Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex. Protein Science, 21(10), 1528-1539.
Niemann, H., Gherardi, E., Bleymüller, W., and Heinz, D. W. (2012). Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex. Protein Science 21, 1528-1539.
Niemann, H., et al., 2012. Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex. Protein Science, 21(10), p 1528-1539.
H. Niemann, et al., “Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex”, Protein Science, vol. 21, 2012, pp. 1528-1539.
Niemann, H., Gherardi, E., Bleymüller, W., Heinz, D.W.: Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex. Protein Science. 21, 1528-1539 (2012).
Niemann, Hartmut, Gherardi, Ermanno, Bleymüller, Willem, and Heinz, Dirk W. “Engineered variants of InlB with an additional leucine-rich repeat discriminate between physiologically relevant and packing contacts in crystal structures of the InlB:MET complex”. Protein Science 21.10 (2012): 1528-1539.
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51 References

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MolProbity: all-atom structure validation for macromolecular crystallography
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