While the KinCon reporter approach offers valuable insights into kinase conformational dynamics, there are some limitations and caveats that should be considered:
Overexpression Artifacts: High levels of exogenous expression of KinCon reporters may lead to non-physiological interactions and conformational changes. Optimizing the expression levels of KinCon reporters to mimic endogenous kinase levels can help mitigate this issue.
Cell Line Specificity: The behavior of KinCon reporters may vary across different cell lines, potentially impacting the reliability and reproducibility of the results. Validating the KinCon technology in multiple cell types can help address this limitation.
Limited Spatial Resolution: The KinCon reporter technology provides information on kinase conformational changes at the whole-cell level, lacking spatial resolution. Combining KinCon reporters with spatially resolved techniques can help overcome this limitation.
Inability to Capture Post-Translational Modifications: KinCon reporters may not capture the effects of post-translational modifications on kinase conformation. Integrating KinCon technology with phospho-specific antibodies or mass spectrometry can provide a more comprehensive view of kinase regulation.
Need for Validation: Validating the KinCon reporter results with orthogonal techniques such as biochemical assays or structural studies is essential to ensure the accuracy and reliability of the findings.