A time-resolved luminescence biosensor assay for anaplastic lymphoma kinase (ALK) activity

Cui, Wei; Parker, Laurie L.

doi:10.1039/c4cc07453j

Cited by 22 publications

(21 citation statements)

References 24 publications

(51 reference statements)

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“…Other phosphotyrosine (pTyr) containing peptides have been developed that bind to Tb(III), using the pTyr residue to sensitise Tb(III) emission. [101][102][103][104][105] An acidic peptide was used, containing a central tyrosine residue, which acts as a specific substrate for spleen tyrosine kinase (Syk). 101,106 The phosphorylated peptide was found to bind Tb(III) ions 5 times more strongly than to the non-phosphorylated analogue, and with a substantial increase in emission intensity.…”

Section: Using Phosphorylation To Affect Ln(iii) Ion Peptide Bindingmentioning

confidence: 99%

Application of lanthanide luminescence in probing enzyme activity

Hewitt

Butler

2018

Chem. Commun.

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Enzymes play critical roles in the regulation of cellular function and are implicated in numerous disease conditions. Reliable and practicable assays are required to study enzyme activity, to facilitate the discovery of inhibitors and activators of enzymes related to disease. In recent years, a variety of enzyme assays have been devised that utilise luminescent lanthanide(iii) complexes, taking advantage of their high detection sensitivities, long luminescence lifetimes, and line-like emission spectra that permit ratiometric and time-resolved analyses. In this Feature article, we focus on recent progress in the development of enzyme activity assays based on lanthanide(iii) luminescence, covering a variety of strategies including Ln(iii)-labelled antibodies and proteins, Ln(iii) ion encapsulation within defined peptide sequences, reactivity-based Ln(iii) probes, and discrete Ln(iii) complexes. Emerging approaches for monitoring enzyme activity are discussed, including the use of anion responsive lanthanide(iii) complexes, capable of molecular recognition and luminescence signalling of polyphosphate anions.

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Section: Using Phosphorylation To Affect Ln(iii) Ion Peptide Bindingmentioning

confidence: 99%

Application of lanthanide luminescence in probing enzyme activity

Hewitt

Butler

2018

Chem. Commun.

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“…Additionally, our panel included the receptor tyrosine kinase ALK, which we previously observed to phosphorylate similar sequences to those phosphorylated by SRC. 41 To ensure each of those recombinant kinases was active, we performed an in vitro kinase assay with several reference peptides that have been previously characterized in our laboratory for the kinases in the panel.…”

Section: In Vitro Characterization Of Fastide Flt3 Specificitymentioning

confidence: 99%

“…The "universal" tyrosine kinase substrate that we previously reported U5 (DEAIYATVA) 34 was the reference peptide chosen for KIT, PDGFRβ and BTK, and an ALK substrate we previously reported (ALAStide) 41,42 was used for ALK ( Figure 5A-F). SFAStide-A (DEDIYEELD) 31 was used as the reference peptide for SRC and LYN kinases.…”

Section: In Vitro Characterization Of Fastide Flt3 Specificitymentioning

confidence: 99%

High-throughput identification of FLT3 wild-type and mutant kinase substrate preferences and application to design of sensitive in vitro kinase assay substrates

Perez

Blankenhorn

Murray

et al. 2018

Preprint

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Acute myeloid leukemia (AML) is an aggressive disease that is characterized by abnormal increase of immature myeloblasts in blood and bone marrow. The FLT3 receptor tyrosine kinase plays an integral role in haematopoiesis, and one third of AML diagnoses exhibit gain-offunction mutations in FLT3, with the juxtamembrane domain internal tandem duplication (ITD) and the kinase domain D835Y variants observed most frequently. Few FLT3 substrates or phosphorylation sites are known, which limits insight into FLT3's substrate preferences and makes assay design particularly challenging. We applied in vitro phosphorylation of a cell lysate digest (adaptation of the Kinase Assay Linked with Phosphoproteomics (KALIP) technique and similar methods) for high-throughput identification of substrates for three FLT3 variants (wildtype, ITD mutant, and D835Y mutant). Incorporation of identified substrate sequences as input into the KINATEST-ID substrate preference analysis and assay development pipeline facilitated the design of several peptide substrates that are phosphorylated efficiently by all three FLT3 kinase variants. These substrates could be used in assays to identify new FLT3 inhibitors that overcome resistant mutations to improve FLT3-positive AML treatment.

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“…These changes include enhancing the Tb 3+ binding affinity, reducing the Tb 3+ chelate hydration number, increasing the Tb 3+ luminescence lifetime, and shifting the excitation wavelength of tyrosine. 21–23 …”

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confidence: 99%

Multicolored, Tb³⁺-Based Antibody-Free Detection of Multiple Tyrosine Kinase Activities

et al. 2015

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Kinase signaling is a major mechanism driving many cancers. While many inhibitors have been developed and are employed in the clinic, resistance due to crosstalk and pathway reprogramming is an emerging problem. High-throughput assays to detect multiple pathway kinases simultaneously could better model these complex relationships and enable drug development to combat this type of resistance. We developed a strategy to take advantage of time-resolved luminescence of Tb3+-chelated phosphotyrosine-containing peptides, which facilitated efficient energy transfer to small molecule fluorophores conjugated to the peptides to produce orthogonally-colored biosensors for two different kinases. This enabled multiplexed detection with high signal to noise in a high-throughput-compatible format. This proof-of-concept study provides a platform that could be applied to other lanthanide metal and fluorophore combinations to achieve even greater multiplexing without the need for phosphospecific antibodies.

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A time-resolved luminescence biosensor assay for anaplastic lymphoma kinase (ALK) activity

Cited by 22 publications

References 24 publications

Application of lanthanide luminescence in probing enzyme activity

Application of lanthanide luminescence in probing enzyme activity

High-throughput identification of FLT3 wild-type and mutant kinase substrate preferences and application to design of sensitive in vitro kinase assay substrates

Multicolored, Tb³⁺-Based Antibody-Free Detection of Multiple Tyrosine Kinase Activities

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A time-resolved luminescence biosensor assay for anaplastic lymphoma kinase (ALK) activity

Cited by 22 publications

References 24 publications

Application of lanthanide luminescence in probing enzyme activity

Application of lanthanide luminescence in probing enzyme activity

High-throughput identification of FLT3 wild-type and mutant kinase substrate preferences and application to design of sensitive in vitro kinase assay substrates

Multicolored, Tb3+-Based Antibody-Free Detection of Multiple Tyrosine Kinase Activities

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Multicolored, Tb³⁺-Based Antibody-Free Detection of Multiple Tyrosine Kinase Activities