Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution

Published

November 7, 2011

Author(s)

Illarion V. Turko, Natalia Gorlatova, Kinlin L. Chao, Lipika Ray, Rawan Araj, Andrey Galkin, John Moult, Osnat Herzberg

Abstract

Macrophage Stimulating Protein (MSP) is the ligand of RON receptor tyrosine kinase. MSP binding to RON activates a signaling pathway that plays a dual role in inflammation, initially stimulating macrophage chemotaxis and phagocytosis, but also changing macrophage state to downregulate inflammatory response. Genome association mapping identified linkage between Crohn’s disease (CD) and ulcerative colitis (UC) and a non-synonymous single nucleotide polymorphism in the MSP gene, MST1, which generates a mutation in the MSP β-chain, R689C. To determine the molecular mechanism by which R689C MSP might be involved in CD and UC, we have produced and purified the wild-type and mutant MSP β-chains (MSPβ) and compared some of their key biochemical and biophysical properties that might impact the MSP/RON signaling pathway. Mass spectrometry of tryptic fragments showed that the new thiol introduced by the R689C mutation did not form an aberrant disulphide bond. The mutation did not impair the specific Arg383-Val384 peptide bond cleavage by matriptase-1 required for MSP activation. In contrast, differential scanning fluorimetry showed that the thermal stability of the mutant MSPβ was lower than that of wild-type MSPβ by 1.6 °C. Additionally, surface plasmon resonance binding studies showed that the R689C MSPβ affinity towards RON was approximately 10-fold lower than that of the wild-type MSPβ. Together, the studies indicate that the mutation might impair MSP function by reducing its half-life in vivo and its affinity to RON, both of which might down-regulate the MSP/RON signaling pathway.

Citation

PLoS One

Volume

6

Issue

11

Pub Type

Journals

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DOI Link

Keywords

macrophage stimulating protein, mass spectrometry, differential scanning fluorimetry, surface plasmon resonance

Citation

Turko, I. , Gorlatova, N. , Chao, K. , Ray, L. , Araj, R. , Galkin, A. , Moult, J. and Herzberg, O. (2011), Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution, PLoS One, [online], https://doi.org/10.1371/journal.pone.0027269 (Accessed December 13, 2024)