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Identifying Kinase Substrates via a Heavy ATP Kinase Assay and Quantitative Mass Spectrometry

ABSTRACT: Mass spectrometry‐based in vitro kinase screens play an essential role in the discovery of kinase substrates, however, many suffer from biological and technical noise or necessitate genetically‐altered enzymecofactor systems. We describe a method that combines stable γ‐[18O2]‐ATP with classical in vitro kinase assays within a contemporary quantitative proteomic workflow. Our approach improved detection of known substrates of the non‐receptor tyrosine kinase ABL1; and identified potential, new in vitro substrates.

INSTRUMENT(S): LTQ Orbitrap Velos, Q Exactive

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Epithelial Cell, Kidney

DISEASE(S): Acute Leukemia

SUBMITTER: Andre Mueller

LAB HEAD: Keiryn L. Bennett

PROVIDER: PXD002133 | Pride | 2018-10-17

REPOSITORIES: Pride

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Identifying Kinase Substrates via a Heavy ATP Kinase Assay and Quantitative Mass Spectrometry.

Müller André C AC Giambruno Roberto R Weißer Juliane J Májek Peter P Hofer Alexandre A Bigenzahn Johannes W JW Superti-Furga Giulio G Jessen Henning J HJ Bennett Keiryn L KL

Scientific reports 20160627

Mass spectrometry-based in vitro kinase screens play an essential role in the discovery of kinase substrates, however, many suffer from biological and technical noise or necessitate genetically-altered enzyme-cofactor systems. We describe a method that combines stable γ-[(18)O2]-ATP with classical in vitro kinase assays within a contemporary quantitative proteomic workflow. Our approach improved detection of known substrates of the non-receptor tyrosine kinase ABL1; and identified potential, new ...[more]

PMID: 27346722

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