Cooperativity between verapamil and ATP bound to the efflux transporter P-glycoprotein

Ledwitch, Kaitlyn; Gibbs, Morgan E.; Barnes, Robert W.; Roberts, Arthur G.

doi:10.1016/j.bcp.2016.08.013

Cited by 50 publications

(78 citation statements)

References 100 publications

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“…4. To obtain microscopic kinetic parameters, such as V max s and K d s, experimentally determined data were fit with COPASI, as previously described (60, 63), using the kinetic model in Fig. 4.…”

Section: Resultsmentioning

confidence: 99%

“…Data were fitted by using Michaelis‐Menten equation using Igor Pro 6.2 (58–60). Monophasic ATP hydrolysis kinetics were fit to a modified Michaelis‐Menten equation (58–60):

v = \frac{V_{\max} [L]}{K_{normalM} + [L]} + v_{basal}

Section: Methodsmentioning

confidence: 99%

“…Monophasic ATP hydrolysis kinetics were fit to a modified Michaelis‐Menten equation (58–60):

v = \frac{V_{\max} [L]}{K_{normalM} + [L]} + v_{basal}

where v is the ATP hydrolysis rate, V max is the maximum ATP hydrolysis rate, v basal is the basal ATP hydrolysis rate, K M is the Michaelis‐Menten constant, and L is the transported substrate. Biphasic ATP hydrolysis kinetics were fit to modified substrate inhibition equation (58–60):

v = \frac{V_{\max}}{1 + \frac{K_{normalM}}{[L]} + \frac{[L]}{K_{normali}}} + v_{basal}

where K I is the inhibitory constant. Monophasic transport kinetics were fit to another variation of the Michaelis‐Menten equation (58, 59):

t = \frac{T_{\max} [L]}{K_{normalM} + [L]}

where t is the apparent transport rate and T max is the maximum transport rate.…”

Section: Methodsmentioning

confidence: 99%

“…As a result, a variety of advanced software modeling packages have been developed to fit arbitrary kinetic models, including the free Complex Pathway Simulator (COPASI) (62) and the proprietary Berkeley Madonna (University of California, Berkeley, Berkeley, CA, USA). To estimate discrete microscopic kinetic parameters, such as V max s and K d s (dissociation constant), transport and DMRP‐mediated ATP hydrolysis in this study were fit to kinetic models using the evolutionary algorithm in the COPASI software as previously described (60, 62, 63). This software can fit both kinetic and steady‐state curves with an indeterminate number of independent fitting parameters (63).…”

Section: Methodsmentioning

confidence: 99%

“…Data were fitted by using Michaelis-Menten equation using Igor Pro 6.2 (58)(59)(60). Monophasic ATP hydrolysis kinetics were fit to a modified Michaelis-Menten equation (58)(59)(60):…”

Section: Calculation Of Kinetic Parameters Of Atpase and Vesicular Trmentioning

confidence: 99%

See 4 more Smart Citations

Boosted coupling of ATP hydrolysis to substrate transport upon cooperative estradiol‐17‐β‐D‐glucuronide binding in aDrosophilaATP binding cassette type‐C transporter

Karasik

Ledwitch²,

Arányi

et al. 2018

FASEB j.

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ATP binding cassette type-C (ABCC) transporters move molecules across cell membranes upon hydrolysis of ATP; however, their coupling of ATP hydrolysis to substrate transport remains elusive. Drosophila multidrug resistance-associated protein (DMRP) is the functional ortholog of human long ABCC transporters, with similar substrate and inhibitor specificity, but higher activity. Exploiting its high activity, we kinetically dissected the catalytic mechanism of DMRP by using E-d-glucuronide (EG), the physiologic substrate of human ABCC. We examined the DMRP-mediated interdependence of ATP and EG in biochemical assays. We observed EG-dependent ATPase activity to be biphasic at subsaturating ATP concentrations, which implies at least 2 EG binding sites on DMRP. Furthermore, transport measurements indicated strong nonreciprocal cooperativity between ATP and EG. In addition to confirming these findings, our kinetic modeling with the Complex Pathway Simulator indicated a 10-fold decrease in the EG-mediated activation of ATP hydrolysis upon saturation of the second EG binding site. Surprisingly, the binding of the second EG allowed for substrate transport with a constant rate, which tightly coupled ATP hydrolysis to transport. In summary, we show that the second EG binding-similar to human ABCC2-allosterically stimulates transport activity of DMRP. Our data suggest that this is achieved by a significant increase in the coupling of ATP hydrolysis to transport.-Karasik, A., Ledwitch, K. V., Arányi, T., Váradi, A., Roberts, A., Szeri, F. Boosted coupling of ATP hydrolysis to substrate transport upon cooperative estradiol-17-β-D-glucuronide binding in a Drosophila ATP binding cassette type-C transporter.

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Section: Resultsmentioning

confidence: 99%

“…Data were fitted by using Michaelis‐Menten equation using Igor Pro 6.2 (58–60). Monophasic ATP hydrolysis kinetics were fit to a modified Michaelis‐Menten equation (58–60):

v = \frac{V_{\max} [L]}{K_{normalM} + [L]} + v_{basal}

Section: Methodsmentioning

confidence: 99%

“…Monophasic ATP hydrolysis kinetics were fit to a modified Michaelis‐Menten equation (58–60):

v = \frac{V_{\max} [L]}{K_{normalM} + [L]} + v_{basal}

v = \frac{V_{\max}}{1 + \frac{K_{normalM}}{[L]} + \frac{[L]}{K_{normali}}} + v_{basal}

where K I is the inhibitory constant. Monophasic transport kinetics were fit to another variation of the Michaelis‐Menten equation (58, 59):

t = \frac{T_{\max} [L]}{K_{normalM} + [L]}

where t is the apparent transport rate and T max is the maximum transport rate.…”

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

“…Data were fitted by using Michaelis-Menten equation using Igor Pro 6.2 (58)(59)(60). Monophasic ATP hydrolysis kinetics were fit to a modified Michaelis-Menten equation (58)(59)(60):…”

Section: Calculation Of Kinetic Parameters Of Atpase and Vesicular Trmentioning

confidence: 99%

See 3 more Smart Citations

Boosted coupling of ATP hydrolysis to substrate transport upon cooperative estradiol‐17‐β‐D‐glucuronide binding in aDrosophilaATP binding cassette type‐C transporter

Karasik

Ledwitch²,

Arányi

et al. 2018

FASEB j.

View full text Add to dashboard Cite

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Conformational changes in the nucleotide‐binding domains of P‐glycoprotein induced by ATP hydrolysis

2020

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P‐glycoprotein (Pgp) is a member of the ABC transporter superfamily with high physiological importance. Pgp nucleotide‐binding domains (NBDs) drive the transport cycle through ATP binding and hydrolysis. We use molecular dynamics simulations to investigate the ATP hydrolysis‐induced conformational changes in NBDs. Five systems, including all possible ATP/ADP combinations in the NBDs and the APO system, are simulated. ATP/ADP exchange induces conformational changes mostly within the conserved signature motif of the NBDs, resulting in relative orientational changes in the NBDs. Nucleotide removal leads to additional orientational changes in the NBDs, allowing their dissociation. Furthermore, we capture putative hydrolysis‐competent configurations in which the conserved glutamate in the Walker‐B motif acts as a catalytic base capturing a water molecule likely initiating ATP hydrolysis.

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Sensitizing multidrug-resistant leukemia cells to common cytostatics by an aluminium-salen complex that has high-apoptotic effects in leukemia, lymphoma and mamma carcinoma cells

et al. 2021

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Cooperativity between verapamil and ATP bound to the efflux transporter P-glycoprotein

Cited by 50 publications

References 100 publications

Boosted coupling of ATP hydrolysis to substrate transport upon cooperative estradiol‐17‐β‐D‐glucuronide binding in aDrosophilaATP binding cassette type‐C transporter

Boosted coupling of ATP hydrolysis to substrate transport upon cooperative estradiol‐17‐β‐D‐glucuronide binding in aDrosophilaATP binding cassette type‐C transporter

Conformational changes in the nucleotide‐binding domains of P‐glycoprotein induced by ATP hydrolysis

Sensitizing multidrug-resistant leukemia cells to common cytostatics by an aluminium-salen complex that has high-apoptotic effects in leukemia, lymphoma and mamma carcinoma cells

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