Auto-thiophosphorylation activity of Src tyrosine kinase

Cabail, Maria Zulema; Chen, Emily I.; Koller, Antonius; Miller, W. Todd

doi:10.1186/s12858-016-0071-z

Cited by 5 publications

(3 citation statements)

References 33 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…5A). A number of cellular kinases have affinity for ATP-γS (Cabail et al, 2016; Ikeda et al, 1991) and can accept it as an alternative to ATP from which to transfer the γ-thiol to endogenous target proteins. We therefore used the exogenous addition of ATP-γS to the medium to directly assess whether fission yeast assimilates environmental ATP.…”

Section: Resultsmentioning

confidence: 99%

Import of extracellular ATP in yeast and man modulates AMPK and TORC1 signalling

Forte

Davie

Lie

et al. 2019

Journal of Cell Science

View full text Add to dashboard Cite

AMP-activated kinase (AMPK) and target of rapamycin (TOR) signalling coordinate cell growth, proliferation, metabolism and cell survival with the nutrient environment of cells. The poor vasculature and nutritional stress experienced by cells in solid tumours raises the question: how do they assimilate sufficient nutrients to survive? Here, we show that human and fission yeast cells import ATP and AMP from their external environment to regulate AMPK and TOR signalling. Exposure of fission yeast ( Schizosaccharomyces pombe ) and human cells to external AMP impeded cell growth; however, in yeast this restraining impact required AMPK. In contrast, external ATP rescued the growth defect of yeast mutants with reduced TORC1 signalling; furthermore, exogenous ATP transiently enhanced TORC1 signalling in both yeast and human cell lines. Addition of the PANX1 channel inhibitor probenecid blocked ATP import into human cell lines suggesting that this channel may be responsible for both ATP release and uptake in mammals. In light of these findings, it is possible that the higher extracellular ATP concentration reported in solid tumours is both scavenged and recognized as an additional energy source beneficial for cell growth.

show abstract

Section: Resultsmentioning

confidence: 99%

Import of extracellular ATP in yeast and man modulates AMPK and TORC1 signalling

Forte

Davie

Lie

et al. 2019

Journal of Cell Science

View full text Add to dashboard Cite

show abstract

“…Selections employing ATP-γS as a co-substrate with subsequent labeling of phosphothiolates were additionally explored but were met with several problems [7,10]. ATP-γS is a non-natural and poor substrate for c-Src [48]. Commercially available ATP-γS is significantly contaminated with high levels of inhibitory ADP [49].…”

Section: Resultsmentioning

confidence: 99%

Application of a Substrate-Mediated Selection with c-Src Tyrosine Kinase to a DNA-Encoded Chemical Library

et al. 2019

View full text Add to dashboard Cite

As aberrant activity of protein kinases is observed in many disease states, these enzymes are common targets for therapeutics and detection of activity levels. The development of non-natural protein kinase substrates offers an approach to protein substrate competitive inhibitors, a class of kinase inhibitors with promise for improved specificity. Also, kinase activity detection approaches would benefit from substrates with improved activity and specificity. Here, we apply a substrate-mediated selection to a peptidomimetic DNA-encoded chemical library for enrichment of molecules that can be phosphorylated by the protein tyrosine kinase, c-Src. Several substrates were identified and characterized for activity. A lead compound (SrcDEL10) showed both the ability to serve as a substrate and to promote ATP hydrolysis by the kinase. In inhibition assays, compounds displayed IC50′s ranging from of 8–100 µM. NMR analysis of SrcDEL10 bound to the c-Src:ATP complex was conducted to characterize the binding mode. An ester derivative of the lead compound demonstrated cellular activity with inhibition of Src-dependent signaling in cell culture. Together, the results show the potential for substrate-mediated selections of DNA-encoded libraries to discover molecules with functions other than simple protein binding and offer a new discovery method for development of synthetic tyrosine kinase substrates.

show abstract

“…More importantly, many kinases show poor catalytic efficiency with ATPγ‐S, thereby somewhat limiting the applicability of this approach across the kinome (Table ). However, for many kinases, substituting MgCl 2 (or MnCl 2 ) with NiCl 2 in the kinase buffer remarkably enhances the K cat of kinases with thio‐ATP , which could be employed with capture and release ASKA methodology. Despite these limitations, thio‐ATP shows very low background signal from endogenous kinases, making this method a popular choice for kinase–substrate identification on a global scale.…”

Section: Direct Tagging Of Kinase Substratesmentioning

confidence: 99%

The significant others: Global search for direct kinase substrates using chemical approaches

Shah

Kim

2019

IUBMB Life

View full text Add to dashboard Cite

Protein kinases function as key signaling hubs in the intricate network of biochemical signaling processes in the living cell. More than two‐thirds of the human proteome is estimated to be phosphorylated at ~960,000 phosphosites, which makes it challenging to identify the direct contribution of any desired kinase in generating this phosphoproteome. In this review, we discuss some of the methods that have been developed over the years for global identification of kinase substrates. The methods are essentially categorized into two classes, namely, (i) direct tagging of kinase substrates and (ii) indirect phosphoproteomics‐based approaches. We discuss the advantages and limitations entailed to each of the method introduced, with a special emphasis on the analog‐sensitive (as) kinase approach method. © 2019 IUBMB Life, 71(6):721–737, 2019

show abstract

Auto-thiophosphorylation activity of Src tyrosine kinase

Cited by 5 publications

References 33 publications

Import of extracellular ATP in yeast and man modulates AMPK and TORC1 signalling

Import of extracellular ATP in yeast and man modulates AMPK and TORC1 signalling

Application of a Substrate-Mediated Selection with c-Src Tyrosine Kinase to a DNA-Encoded Chemical Library

The significant others: Global search for direct kinase substrates using chemical approaches

Contact Info

Product

Resources

About