The human P-glycoprotein (Pgp) 3 functions as an ATP-driven drug transporter conferring multidrug resistance in cancer cells and restricting bioavailability of many antimicrobial and anticancer agents (1, 2). It is a 1280-amino acid integral membrane protein of the ATP-binding cassette (ABC) transporter family (3, 4), with two highly homologous halves, each containing a hydrophobic transmembrane domain and a relatively hydrophilic cytosolic domain. Each hydrophobic domain contains six putative transmembrane helices that, in conjunction with the transmembrane regions of the other half, form the drug-translocating pathway (substrate site) across the lipid bilayer (1, 5). The cytosolic domains each contain three consensus sequences (3) that together contribute to the formation of two ATP binding/hydrolysis sites (ATP sites) (6).Both ATP sites are essential for the drug transport function of Pgp (7-10) but are believed to hydrolyze ATP in an alternate sequence (11-13). Pgp possesses a basal rate of ATP hydrolysis that is stimulated upon interaction with many transport substrates as well as with several Pgp modulators (14). Binding and hydrolysis of ATP induce a conformational change in that is coupled to substrate translocation across the lipid bilayer and its subsequent dissociation (18 -24). The two ATP sites of Pgp were believed to be functionally equivalent with drug translocation and regeneration of the transporter directly coupled to a single round of ATP binding or/and hydrolysis (12,25,26). However, evidence suggests that although the two ATP sites assume similar structural conformation, they have distinct functional roles within a single catalytic turnover of (18,19), whereas hydrolysis by the other resets the transporter for the subsequent round of transport activity (27,28). Vanadate (Vi), a phosphate analog, replaces phosphate at the catalytic site (30) when present during ATP hydrolysis. Due to its low rate of dissociation, vanadate stabilizes Pgp in a catalytic intermediate (mimicking a conformational state) that immediately follows the first ATP hydrolytic event (11,18,19,22,23 (18,19). This is accompanied by an experimentally detectable conformational change (15,17). Chemical cross-linking with thiol reagents and cysteine-scanning mutagenesis revealed increased accessibility of the drug-translocating pathway from the extracellular side of the lipid bilayer due to vanadate trapping (22).Interestingly, the [ 125 I]IAAP-binding site once transformed to its low affinity state remains unaltered even after dissociation of vanadate and
The human P-glycoprotein (Pgp, ABCB1) is an ATP-dependent efflux pump for structurally unrelated hydrophobic compounds, conferring simultaneous resistance to and restricting bioavailability of several anticancer and antimicrobial agents. Drug transport by Pgp requires a coordinated communication between its substrate binding/translocating pathway (substrate site) and the nucleotide binding domains (NBDs or ATP sites). In this study, we demonstrate that certain thioxanthene-based Pgp modulators, such as cis-(Z)-flupentixol and its closely related analogues, effectively disrupt molecular cross talk between the substrate, and the ATP, sites without affecting the basic functional aspects of the two domains, such as substrate recognition, binding, and hydrolysis of ATP and dissociation of ADP following ATP hydrolysis. The allosteric modulator cis-(Z)-flupentixol has no effect on [alpha-(32)P]-8-azido-ATP binding to Pgp under nonhydrolytic conditions or on the K(m) for ATP during ATP hydrolysis. Both hydrolysis of ATP and vanadate-induced [alpha-(32)P]-8-azido-ADP trapping (following [alpha-(32)P]-8-azido-ATP breakdown) by Pgp are stimulated by the modulator. However, the ability of Pgp substrates (such as prazosin) to stimulate ATP hydrolysis and facilitate vanadate-induced trapping of [alpha-(32)P]-8-azido-ADP is substantially affected in the presence of cis-(Z)-flupentixol. Substrate recognition by Pgp as determined by [(125)I]iodoarylazidoprazosin ([(125)I]IAAP) binding both in the presence and in the absence of ATP is facilitated by the modulator, whereas substrate dissociation in response to vanadate trapping is considerably affected in its presence. In the Pgp F983A mutant, which is impaired in modulation by cis-(Z)-flupentixol, the modulator has a minimal effect on substrate-stimulated ATP hydrolysis as well as on substrate dissociation coupled to vanadate trapping. Finally, cis-(Z)-flupentixol has no effect on dissociation of [alpha-(32)P]-8-azido-ADP (or ADP) from vanadate-trapped Pgp, which is essential for subsequent rounds of ATP hydrolysis. Taken together, our results demonstrate a distinct mechanism of Pgp modulation that involves allosteric disruption of molecular cross talk between the substrate, and the ATP, sites without any direct interference with their individual functions.
This scoping study investigated how evidence-based practice (EBP) principles are taught in Australian speech-language pathology (SLP) teaching and learning contexts. It explored how Australian SLP university programs: (1) facilitate student learning about the principles of EBP in academic and clinical settings, and (2) self-evaluate their curricula in relation to EBP. The research involved two surveys. Survey 1 respondents were 131 academic staff, program coordinators, and on-campus and off-campus clinical educators. This survey gathered information about EBP teaching and learning in SLP programs as well as future EBP curriculum plans. Survey 2 investigated how clinical educators incorporated EBP into the way they taught clinical decision-making to students. Surveys responses from 85 clinical educators were analysed using descriptive and non-parametric statistics and thematic grouping of open-ended qualitative responses. Both surveys revealed strengths and gaps in integrating EBP into Australian SLP curricula. Perceived strengths were that respondents were positive about EBP, most had EBP training and access to EBP resources. The perceived gaps included the academic staff's perceptions of students' understanding and application of EBP, respondents' understanding of research methodologies, communication and collaboration between academic staff and clinical educators, and a lack of explicit discussion by clinical educators and students of EBP in relation to clients.
Oral lichen planus (OLP) is a stress induced inflammatory condition with malignant potency. The mdr1 (multidrug resistance) is a stress gene overexpressed in cancerous conditions and its translated form, the p-glycoprotein efflux transporter is usually overexpressed with chemotherapy, leading to chemoresistance. OLP, a lesion with carcinogenic potency, is broadly classified into the asymptomatic reticular form and the aggressive erosive form. The objective of the study was to verify the expression level of p-glycoprotein in antifungaltreated and untreated reticular OLP, in untreated erosive OLP and erosive OLP patients treated with corticosteroid. Semi-quantitative reverse transcriptase polymerase chain reaction (SQ-RTPCR) and ELISA were performed on biopsy tissue samples to evaluate the mdr1 mRNA and protein expression of p-glycoprotein, respectively. The present study shows for the first time that mdr1 mRNA as well as its translated form p-glycoprotein are overexpressed in OLP subjects compared to healthy individuals. This overexpression is significantly higher in erosive than in reticular OLP patients, further confirming that the erosive form has higher risk for multidrug resistance. A higher expression is also observed in corticosteroid-treated erosive cases than similar untreated ones. The gradation of expression is in conformity with severity of the disease.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.