Structure-Function Analysis of the Non-Muscle Myosin Light Chain Kinase (nmMLCK) Isoform by NMR Spectroscopy and Molecular Modeling: Influence of MYLK Variants.

TitleStructure-Function Analysis of the Non-Muscle Myosin Light Chain Kinase (nmMLCK) Isoform by NMR Spectroscopy and Molecular Modeling: Influence of MYLK Variants.
Publication TypeJournal Article
Year of Publication2015
AuthorsShen K, Ramirez B, Mapes B, Shen GR, Gokhale V, Brown ME, Santarsiero B, Ishii Y, Dudek SM, Wang T, Garcia JGN
JournalPLoS One
Volume10
Issue6
Paginatione0130515
Date Published2015
ISSN Number1932-6203
KeywordsAfrican Continental Ancestry Group, Humans, Lung Diseases, Magnetic Resonance Spectroscopy, Models, Molecular, Myosin-Light-Chain Kinase, Nuclear Magnetic Resonance, Biomolecular, Phosphorylation, Polymorphism, Single Nucleotide, Protein Isoforms, Structure-Activity Relationship
Abstract

<p>The MYLK gene encodes the multifunctional enzyme, myosin light chain kinase (MLCK), involved in isoform-specific non-muscle and smooth muscle contraction and regulation of vascular permeability during inflammation. Three MYLK SNPs (P21H, S147P, V261A) alter the N-terminal amino acid sequence of the non-muscle isoform of MLCK (nmMLCK) and are highly associated with susceptibility to acute lung injury (ALI) and asthma, especially in individuals of African descent. To understand the functional effects of SNP associations, we examined the N-terminal segments of nmMLCK by 1H-15N heteronuclear single quantum correlation (HSQC) spectroscopy, a 2-D NMR technique, and by in silico molecular modeling. Both NMR analysis and molecular modeling indicated SNP localization to loops that connect the immunoglobulin-like domains of nmMLCK, consistent with minimal structural changes evoked by these SNPs. Molecular modeling analysis identified protein-protein interaction motifs adversely affected by these MYLK SNPs including binding by the scaffold protein 14-3-3, results confirmed by immunoprecipitation and western blot studies. These structure-function studies suggest novel mechanisms for nmMLCK regulation, which may confirm MYLK as a candidate gene in inflammatory lung disease and advance knowledge of the genetic underpinning of lung-related health disparities.</p>

DOI10.1371/journal.pone.0130515
Alternate JournalPLoS ONE
PubMed ID26111161
PubMed Central IDPMC4482139
Grant ListP01 HL058064 / HL / NHLBI NIH HHS / United States
R01 HL091889 / HL / NHLBI NIH HHS / United States
HL058064 / HL / NHLBI NIH HHS / United States
HL091889 / HL / NHLBI NIH HHS / United States