Transportomics: screening for substrates of ABC transporters in body fluids using vesicular transport assays
2012
Krumpochova, P. | Sapthu, S. | Brouwers, J.F. | Haas, de, M. | Vos, de, R.C.H. | Borst, P. | Wetering, van de, K.
The ATP-binding cassette (ABC) genes encode the largest family of transmembrane proteins. ABC transporters translocate a wide variety of substrates across membranes, but their physiological function is often incompletely understood. We describe a new method to study the substrate spectrum of ABC transporters: We incubate extracts of mouse urine with membrane vesicles prepared from Spodoptera frugiperda Sf9 insect cells overproducing an ABC transporter and determine the compounds transported into the vesicles by LC/MS-based metabolomics. We illustrate the power of this simple “transportomics” approach using ABCC2, a protein present at sites of uptake and elimination. We identified many new substrates of ABCC2 in urine. These included glucuronides of plant-derived xenobiotics, a class of compounds to which humans are exposed on a daily basis. Moreover, we show that the excretion of these compounds in vivo depends on ABCC2: compared to wild-type mice, the urinary excretion of several glucuronides was increased up to 20-fold in Abcc2-/- mice. Transportomics has broad applicability, as it is not restricted to urine and can be applied to other ATP-dependent transport proteins as well.—Krumpochova, P., Sapthu, S., Brouwers, J. F., de Haas, M., de Vos, R., Borst, P., van de Wetering, K. Transportomics: screening for substrates of ABC transporters in body fluids using vesicular transport assays
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