Reprogramming fatty acyl specificity of lipid kinases via C1 domain engineering
Autor: | Timothy B. Ware, Andreas Gahlmann, Thurl E. Harris, Kwon-Sik Park, Mingxing Zhang, Mitchell E. Granade, Kee-Beom Kim, Ku-Lung Hsu, Caroline E. Franks |
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Rok vydání: | 2019 |
Předmět: |
Proteomics
Diacylglycerol Kinase Protein domain Phosphatidic Acids Protein Engineering Gene Expression Regulation Enzymologic Article Substrate Specificity 03 medical and health sciences Protein Domains Tandem Mass Spectrometry Catalytic Domain Animals Humans Metabolomics Chemoproteomics Molecular Biology 030304 developmental biology C1 domain Diacylglycerol kinase 0303 health sciences Kinase Chemistry 030302 biochemistry & molecular biology Cell Biology Protein engineering Rats HEK293 Cells Biochemistry Molecular Probes lipids (amino acids peptides and proteins) Signal transduction Chromatography Liquid |
Zdroj: | Nature chemical biology |
ISSN: | 1552-4469 |
Popis: | C1 domains are lipid-binding modules that regulate membrane activation of kinases, nucleotide exchange factors and other C1-containing proteins to trigger signal transduction. Despite annotation of typical C1 domains as diacylglycerol (DAG) and phorbol ester sensors, the function of atypical counterparts remains ill-defined. Here, we assign a key role for atypical C1 domains in mediating DAG fatty acyl specificity of diacylglycerol kinases (DGKs) in live cells. Activity-based proteomics mapped C1 probe binding as a principal differentiator of type 1 DGK active sites that combined with global metabolomics revealed a role for C1s in lipid substrate recognition. Protein engineering by C1 domain swapping demonstrated that exchange of typical and atypical C1s is functionally tolerated and can directly program DAG fatty acyl specificity of type 1 DGKs. Collectively, we describe a protein engineering strategy for studying metabolic specificity of lipid kinases to assign a role for atypical C1 domains in cell metabolism. A chemoproteomics and protein engineering strategy to investigate metabolic activity of lipid kinases led to assignment of a key role for atypical C1 domains in directing fatty acyl specificity of diacylglycerol kinase function in live cells. |
Databáze: | OpenAIRE |
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