Deconstructing Lipid Kinase Inhibitors by Chemical Proteomics

Rebecca L. McCloud, Caroline E. Franks, Sean T. Campbell, Benjamin W. Purow, Thurl E. Harris, Ku Lung Hsu

Research output: Contribution to journalArticle

7 Citations (Scopus)

Abstract

Diacylglycerol kinases (DGKs) regulate lipid metabolism and cell signaling through ATP-dependent phosphorylation of diacylglycerol to biosynthesize phosphatidic acid. Selective chemical probes for studying DGKs are currently lacking and are needed to annotate isoform-specific functions of these elusive lipid kinases. Previously, we explored fragment-based approaches to discover a core fragment of DGK-α (DGKα) inhibitors responsible for selective binding to the DGKα active site. Here, we utilize quantitative chemical proteomics to deconstruct widely used DGKα inhibitors to identify structural regions mediating off-target activity. We tested the activity of a fragment (RLM001) derived from a nucleotide-like region found in the DGKα inhibitors R59022 and ritanserin and discovered that RLM001 mimics ATP in its ability to broadly compete at ATP-binding sites of DGKα as well as >60 native ATP-binding proteins (kinases and ATPases) detected in cell proteomes. Equipotent inhibition of activity-based probe labeling by RLM001 supports a contiguous ligand-binding site composed of C1, DAGKc, and DAGKa domains in the DGKα active site. Given the lack of available crystal structures of DGKs, our studies highlight the utility of chemical proteomics in revealing active-site features of lipid kinases to enable development of inhibitors with enhanced selectivity against the human proteome.

Original languageEnglish (US)
Pages (from-to)231-236
Number of pages6
JournalBiochemistry
Volume57
Issue number2
DOIs
StatePublished - Jan 16 2018
Externally publishedYes

Fingerprint

Diacylglycerol Kinase
Proteomics
Phosphotransferases
Lipids
Adenosine Triphosphate
Catalytic Domain
Proteome
R 59022
Binding Sites
Ritanserin
Cell signaling
Phosphatidic Acids
Phosphorylation
Diglycerides
Lipid Metabolism
Protein Kinases
Labeling
Adenosine Triphosphatases
Carrier Proteins
Protein Isoforms

ASJC Scopus subject areas

  • Biochemistry

Cite this

McCloud, R. L., Franks, C. E., Campbell, S. T., Purow, B. W., Harris, T. E., & Hsu, K. L. (2018). Deconstructing Lipid Kinase Inhibitors by Chemical Proteomics. Biochemistry, 57(2), 231-236. https://doi.org/10.1021/acs.biochem.7b00962

Deconstructing Lipid Kinase Inhibitors by Chemical Proteomics. / McCloud, Rebecca L.; Franks, Caroline E.; Campbell, Sean T.; Purow, Benjamin W.; Harris, Thurl E.; Hsu, Ku Lung.

In: Biochemistry, Vol. 57, No. 2, 16.01.2018, p. 231-236.

Research output: Contribution to journalArticle

McCloud, RL, Franks, CE, Campbell, ST, Purow, BW, Harris, TE & Hsu, KL 2018, 'Deconstructing Lipid Kinase Inhibitors by Chemical Proteomics', Biochemistry, vol. 57, no. 2, pp. 231-236. https://doi.org/10.1021/acs.biochem.7b00962
McCloud, Rebecca L. ; Franks, Caroline E. ; Campbell, Sean T. ; Purow, Benjamin W. ; Harris, Thurl E. ; Hsu, Ku Lung. / Deconstructing Lipid Kinase Inhibitors by Chemical Proteomics. In: Biochemistry. 2018 ; Vol. 57, No. 2. pp. 231-236.
@article{616a7795378e4f39b7f2e7e6a4745f1b,
title = "Deconstructing Lipid Kinase Inhibitors by Chemical Proteomics",
abstract = "Diacylglycerol kinases (DGKs) regulate lipid metabolism and cell signaling through ATP-dependent phosphorylation of diacylglycerol to biosynthesize phosphatidic acid. Selective chemical probes for studying DGKs are currently lacking and are needed to annotate isoform-specific functions of these elusive lipid kinases. Previously, we explored fragment-based approaches to discover a core fragment of DGK-α (DGKα) inhibitors responsible for selective binding to the DGKα active site. Here, we utilize quantitative chemical proteomics to deconstruct widely used DGKα inhibitors to identify structural regions mediating off-target activity. We tested the activity of a fragment (RLM001) derived from a nucleotide-like region found in the DGKα inhibitors R59022 and ritanserin and discovered that RLM001 mimics ATP in its ability to broadly compete at ATP-binding sites of DGKα as well as >60 native ATP-binding proteins (kinases and ATPases) detected in cell proteomes. Equipotent inhibition of activity-based probe labeling by RLM001 supports a contiguous ligand-binding site composed of C1, DAGKc, and DAGKa domains in the DGKα active site. Given the lack of available crystal structures of DGKs, our studies highlight the utility of chemical proteomics in revealing active-site features of lipid kinases to enable development of inhibitors with enhanced selectivity against the human proteome.",
author = "McCloud, {Rebecca L.} and Franks, {Caroline E.} and Campbell, {Sean T.} and Purow, {Benjamin W.} and Harris, {Thurl E.} and Hsu, {Ku Lung}",
year = "2018",
month = "1",
day = "16",
doi = "10.1021/acs.biochem.7b00962",
language = "English (US)",
volume = "57",
pages = "231--236",
journal = "Biochemistry",
issn = "0006-2960",
publisher = "American Chemical Society",
number = "2",

}

TY - JOUR

T1 - Deconstructing Lipid Kinase Inhibitors by Chemical Proteomics

AU - McCloud, Rebecca L.

AU - Franks, Caroline E.

AU - Campbell, Sean T.

AU - Purow, Benjamin W.

AU - Harris, Thurl E.

AU - Hsu, Ku Lung

PY - 2018/1/16

Y1 - 2018/1/16

N2 - Diacylglycerol kinases (DGKs) regulate lipid metabolism and cell signaling through ATP-dependent phosphorylation of diacylglycerol to biosynthesize phosphatidic acid. Selective chemical probes for studying DGKs are currently lacking and are needed to annotate isoform-specific functions of these elusive lipid kinases. Previously, we explored fragment-based approaches to discover a core fragment of DGK-α (DGKα) inhibitors responsible for selective binding to the DGKα active site. Here, we utilize quantitative chemical proteomics to deconstruct widely used DGKα inhibitors to identify structural regions mediating off-target activity. We tested the activity of a fragment (RLM001) derived from a nucleotide-like region found in the DGKα inhibitors R59022 and ritanserin and discovered that RLM001 mimics ATP in its ability to broadly compete at ATP-binding sites of DGKα as well as >60 native ATP-binding proteins (kinases and ATPases) detected in cell proteomes. Equipotent inhibition of activity-based probe labeling by RLM001 supports a contiguous ligand-binding site composed of C1, DAGKc, and DAGKa domains in the DGKα active site. Given the lack of available crystal structures of DGKs, our studies highlight the utility of chemical proteomics in revealing active-site features of lipid kinases to enable development of inhibitors with enhanced selectivity against the human proteome.

AB - Diacylglycerol kinases (DGKs) regulate lipid metabolism and cell signaling through ATP-dependent phosphorylation of diacylglycerol to biosynthesize phosphatidic acid. Selective chemical probes for studying DGKs are currently lacking and are needed to annotate isoform-specific functions of these elusive lipid kinases. Previously, we explored fragment-based approaches to discover a core fragment of DGK-α (DGKα) inhibitors responsible for selective binding to the DGKα active site. Here, we utilize quantitative chemical proteomics to deconstruct widely used DGKα inhibitors to identify structural regions mediating off-target activity. We tested the activity of a fragment (RLM001) derived from a nucleotide-like region found in the DGKα inhibitors R59022 and ritanserin and discovered that RLM001 mimics ATP in its ability to broadly compete at ATP-binding sites of DGKα as well as >60 native ATP-binding proteins (kinases and ATPases) detected in cell proteomes. Equipotent inhibition of activity-based probe labeling by RLM001 supports a contiguous ligand-binding site composed of C1, DAGKc, and DAGKa domains in the DGKα active site. Given the lack of available crystal structures of DGKs, our studies highlight the utility of chemical proteomics in revealing active-site features of lipid kinases to enable development of inhibitors with enhanced selectivity against the human proteome.

UR - http://www.scopus.com/inward/record.url?scp=85041801761&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85041801761&partnerID=8YFLogxK

U2 - 10.1021/acs.biochem.7b00962

DO - 10.1021/acs.biochem.7b00962

M3 - Article

C2 - 29155586

AN - SCOPUS:85041801761

VL - 57

SP - 231

EP - 236

JO - Biochemistry

JF - Biochemistry

SN - 0006-2960

IS - 2

ER -