Our results show that Lck-FRET-Zap70FY and Lck-FRET-ITAM1 exhibited the largest increase in ECFP/FRET emission ratio toward Lck kinase activity, while Lck-FRET-Zap70YY was observed to respond fastest among all the biosensors . All the biosensors responded to both active Src and Fyn with similar magnitude and kinetics as Lck (fig. S1).

Expresses a membrane located FRB domain in mammalian cells (the ECFP is not fluorescent)

We developed a new and sensitive FRET biosensor (ZapLck) to visualize Lck kinase activity with high spatiotemporal resolutions in live cells. ZapLck revealed that 62% of Lck signal was preactivated in T-cells.

2013年2月19日 · We applied a Levenberg-Marquardt nonlinear, least-squares algorithm to dissect the decay kinetics of ECFP fluorescence in the Lck biosensor into individual lifetimes. A three-exponential fit resulted in a precise (χ 2 < 1.3) description of the experimental data and revealed three individual τ values of ECFP within the biosensor of 3.2 ns (τ ...

2015年6月15日 · At around 80% confluence, HeLa Kyoto cells were transfected using Lipofectamine 2000 (Invitrogen) according to the manufacturer’s protocol. 1 μg DNA was transfected per well with 3-fold excess of Lck-ECFP-SNAP compared to EGFP-PLCδ-PH and mRFP-FKBP-5Ptase (3:1:1).

2017年5月24日 · Ectopic overexpression of Lck in Th2 cells results in increased expression of CD4 co-receptor and enhanced S73 phosphorylation of transcription factor c-Jun. Our findings indicate that...

2022年7月22日 · Whereas rCD1-dependent recruitment to the PM or the endomembrane system (anchor proteins Lck-ECFP-SNAP or ECFP-SNAP-CAAX, respectively) was not successful, rCD1 induced the recruitment to nucleoli (anchor protein RPL13-ECFP-SNAP) or the nuclear lamina (anchor protein lamin A-ECFP-SNAP).

We developed a new and sensitive FRET biosensor (ZapLck) to visualize Lck kinase activity with high spatiotemporal resolutions in live cells. ZapLck revealed that 62% of Lck signal was...

2019年6月1日 · Lck plays crucial roles in TCR signaling. We developed a new and sensitive FRET biosensor (ZapLck) to visualize Lck kinase activity with high spatiotemporal resolutions in live cells. ZapLck...

To address this, we introduce an advanced ensemble machine learning technique designed to estimate the binding affinity between molecules and LCK. This comprehensive method includes the generation and selection of molecular fingerprints, the design of the machine learning model, hyperparameter tuning, and a model ensemble.