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Mitnaul, L.J. et al.

Mitnaul, Tian, Burton, Lam, Zhu, Olson, Schneeweis, Zuck, Pandit, Anderson, Maletic, Waddell, Wright, Sparrow, Lund,

Fluorogenic substrates for high-throughput measurements of endothelial lipase activity.

Endothelial lipase ( EL) has been shown to be a critical determinant for high density lipoprotein cholesterol levels in vivo; therefore, assays that measure EL activity have become important for the discovery of small molecule inhibitors that specifically target EL. Here, we describe fluorescent Bodipy-labeled substrates that can be used in homogeneous, ultra-high-throughput kinetic assays that measure EL phospholipase or triglyceride lipase activities. Triton X-100 detergent micelles and synthetic HDL particles containing Bodipy- labeled phospholipid or Bodipy- labeled triglyceride substrates were shown to be catalytic substrates for EL, LPL, and HL. More importantly, only synthetic HDL particles containing Bodipy- labeled triglyceride were ideal substrates for EL, LPL, and HL in the presence of high concentrations of human or mouse serum. These data suggest that substrate presentation is a critical factor when determining EL activity in the presence of serum.[1]

References

Fluorogenic substrates for high-throughput measurements of endothelial lipase activity. Mitnaul, L.J., Tian, J., Burton, C., Lam, M.H., Zhu, Y., Olson, S.H., Schneeweis, J.E., Zuck, P., Pandit, S., Anderson, M., Maletic, M.M., Waddell, S.T., Wright, S.D., Sparrow, C.P., Lund, E.G. J. Lipid Res. (2007) [Pubmed]

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