Photoaffinity probes for the alpha 1-adrenergic receptor and the calcium channel bind to a common domain in P-glycoprotein
P-glycoprotein is a 130-180-kDa integral membrane protein that is overproduced in multidrug-resistant cells. The protein appears to act as an energy-dependent drug efflux pump that has broad specificity for structurally diverse hydrophobic antitumor drugs. Many agents, such as the calcium channel bl...
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Veröffentlicht in: | The Journal of biological chemistry 1990-03, Vol.265 (8), p.4394-4401 |
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Zusammenfassung: | P-glycoprotein is a 130-180-kDa integral membrane protein that is overproduced in multidrug-resistant cells. The protein appears
to act as an energy-dependent drug efflux pump that has broad specificity for structurally diverse hydrophobic antitumor drugs.
Many agents, such as the calcium channel blocker verapamil, reverse multidrug resistance and also interact with P-glycoprotein.
The goal of this work was to determine if a common binding site participates in the transport of antitumor drugs and/or the
reversal of drug resistance. This was done by comparing the peptide maps of P-glycoprotein (encoded by mdr1b) after it was
labeled with a photoactive calcium channel blocker, [3H]azidopine, and a newly identified photoaffinity analog for P-glycoprotein
2-[4-(4-azido-3-[125I]iodobenzoyl) piperazin-1-yl]-4-amino-6,7-dimethoxyquinazoline [( 125I]iodoaryl azidoprazosin). [125I]
Iodoaryl azidoprazosin, which classically has been used to identify the alpha 1-adrenergic receptor, bound to P-glycoprotein
and was preferentially competed by vinblastine greater than actinomycin D greater than doxorubicin greater than colchicine.
Peptide maps derived from P-glycoprotein labeled with [3H]azidopine or [125I]iodoaryl azidoprazosin were identical. After
maximal digestion under conditions for Cleveland mapping, a single major 6-kDa fragment was obtained after digestion with
V8 protease, whereas two major fragments, 6.5 and 5.5 kDa, were detected after digestion with chymotrypsin. The 6.0-kDa V8
fragment and the 6.5-kDa chymotrypsin fragment were both found when P-glycoprotein encoded by mdr1a and mdr1b was compared.
Despite its specific interaction with P-glycoprotein, neither iodoaryl azidoprazosin nor prazosin markedly reversed resistance
compared with verapamil or azidopine. Further, multidrug-resistant cells were 900-fold resistant to vinblastine but only 5-fold
resistant to prazosin. These data demonstrate that structurally diverse reversal and/or antitumor agents are likely to have
differential affinity for a small common domain of P-glycoprotein. |
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ISSN: | 0021-9258 1083-351X |
DOI: | 10.1016/S0021-9258(19)39578-X |