Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution.
High throughput genome wide associations studies (GWAS) are now identifying a large number of genome loci related to risk of common human disease. Each such locus presents a challenge in identifying the relevant underlying mechanism. Here we report the experimental characterization of a proposed cau...
Main Authors: | , , , , , , , |
---|---|
Format: | Article |
Language: | English |
Published: |
Public Library of Science (PLoS)
2011-01-01
|
Series: | PLoS ONE |
Online Access: | http://europepmc.org/articles/PMC3210151?pdf=render |
id |
doaj-867cc164b50444d1a20e2d57b2fcbcdc |
---|---|
record_format |
Article |
spelling |
doaj-867cc164b50444d1a20e2d57b2fcbcdc2020-11-24T20:50:08ZengPublic Library of Science (PLoS)PLoS ONE1932-62032011-01-01611e2726910.1371/journal.pone.0027269Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution.Natalia GorlatovaKinlin ChaoLipika R PalRawan Hanna ArajAndrey GalkinIllarion TurkoJohn MoultOsnat HerzbergHigh throughput genome wide associations studies (GWAS) are now identifying a large number of genome loci related to risk of common human disease. Each such locus presents a challenge in identifying the relevant underlying mechanism. Here we report the experimental characterization of a proposed causal single nucleotide polymorphism (SNP) in a locus related to risk of Crohn's disease and ulcerative colitis. The SNP lies in the MST1 gene encoding Macrophage Stimulating Protein (MSP), and results in an R689C amino acid substitution within the β-chain of MSP (MSPβ). MSP binding to the RON receptor tyrosine kinase activates signaling pathways involved in the inflammatory response. We have purified wild-type and mutant MSPβ proteins and compared biochemical and biophysical properties that might impact the MSP/RON signaling pathway. Surface plasmon resonance (SPR) binding studies showed that MSPβ R689C affinity to RON is approximately 10-fold lower than that of the wild-type MSPβ and differential scanning fluorimetry (DSF) showed that the thermal stability of the mutant MSPβ was slightly lower than that of wild-type MSPβ, by 1.6 K. The substitution was found not to impair the specific Arg483-Val484 peptide bond cleavage by matriptase-1, required for MSP activation, and mass spectrometry of tryptic fragments of the mutated protein showed that the free thiol introduced by the R689C mutation did not form an aberrant disulfide bond. Together, the studies indicate that the missense SNP impairs MSP function by reducing its affinity to RON and perhaps through a secondary effect on in vivo concentration arising from reduced thermodynamic stability, resulting in down-regulation of the MSP/RON signaling pathway.http://europepmc.org/articles/PMC3210151?pdf=render |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Natalia Gorlatova Kinlin Chao Lipika R Pal Rawan Hanna Araj Andrey Galkin Illarion Turko John Moult Osnat Herzberg |
spellingShingle |
Natalia Gorlatova Kinlin Chao Lipika R Pal Rawan Hanna Araj Andrey Galkin Illarion Turko John Moult Osnat Herzberg Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution. PLoS ONE |
author_facet |
Natalia Gorlatova Kinlin Chao Lipika R Pal Rawan Hanna Araj Andrey Galkin Illarion Turko John Moult Osnat Herzberg |
author_sort |
Natalia Gorlatova |
title |
Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution. |
title_short |
Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution. |
title_full |
Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution. |
title_fullStr |
Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution. |
title_full_unstemmed |
Protein characterization of a candidate mechanism SNP for Crohn's disease: the macrophage stimulating protein R689C substitution. |
title_sort |
protein characterization of a candidate mechanism snp for crohn's disease: the macrophage stimulating protein r689c substitution. |
publisher |
Public Library of Science (PLoS) |
series |
PLoS ONE |
issn |
1932-6203 |
publishDate |
2011-01-01 |
description |
High throughput genome wide associations studies (GWAS) are now identifying a large number of genome loci related to risk of common human disease. Each such locus presents a challenge in identifying the relevant underlying mechanism. Here we report the experimental characterization of a proposed causal single nucleotide polymorphism (SNP) in a locus related to risk of Crohn's disease and ulcerative colitis. The SNP lies in the MST1 gene encoding Macrophage Stimulating Protein (MSP), and results in an R689C amino acid substitution within the β-chain of MSP (MSPβ). MSP binding to the RON receptor tyrosine kinase activates signaling pathways involved in the inflammatory response. We have purified wild-type and mutant MSPβ proteins and compared biochemical and biophysical properties that might impact the MSP/RON signaling pathway. Surface plasmon resonance (SPR) binding studies showed that MSPβ R689C affinity to RON is approximately 10-fold lower than that of the wild-type MSPβ and differential scanning fluorimetry (DSF) showed that the thermal stability of the mutant MSPβ was slightly lower than that of wild-type MSPβ, by 1.6 K. The substitution was found not to impair the specific Arg483-Val484 peptide bond cleavage by matriptase-1, required for MSP activation, and mass spectrometry of tryptic fragments of the mutated protein showed that the free thiol introduced by the R689C mutation did not form an aberrant disulfide bond. Together, the studies indicate that the missense SNP impairs MSP function by reducing its affinity to RON and perhaps through a secondary effect on in vivo concentration arising from reduced thermodynamic stability, resulting in down-regulation of the MSP/RON signaling pathway. |
url |
http://europepmc.org/articles/PMC3210151?pdf=render |
work_keys_str_mv |
AT nataliagorlatova proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution AT kinlinchao proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution AT lipikarpal proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution AT rawanhannaaraj proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution AT andreygalkin proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution AT illarionturko proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution AT johnmoult proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution AT osnatherzberg proteincharacterizationofacandidatemechanismsnpforcrohnsdiseasethemacrophagestimulatingproteinr689csubstitution |
_version_ |
1716804526215790592 |