Binding site of ABC transporter homology models confirmed by ABCB1 crystal structure

The human ATP-binding cassette (ABC) transporters ABCB1, ABCC4 and ABCC5 are involved in resistance to chemotherapeutic agents. Here we present molecular models of ABCB1, ABCC4 and ABCC5 by homology based on a wide open inward-facing conformation of Escherichia coli MsbA, which were constructed in order to elucidate differences in the electrostatic and molecular features of their drug recognition conformations. As a quality assurance of the methodology, the ABCB1 model was compared to an ABCB1 X-ray crystal structure, and with published crosslinking and site directed mutagenesis data of ABCB1. Amino acids Ile306 (TMH5), Ile340 (TMH6), Phe343 (TMH6), Phe728 (TMH7), and Val982 (TMH12), form a putative substrate recognition site in the ABCB1 model, which is confirmed by both the ABCB1 X-ray crystal structure and the sitedirected mutagenesis studies. The ABCB1, ABCC4 and ABCC5 models display distinct differences in the electrostatic properties of their drug recognition sites

Molecular model of the outward facing state of the human P-glycoprotein (ABCB1), and comparison to a model of the human MRP5 (ABCC5)

<p>Abstract</p> <p>Background</p> <p>Multidrug resistance is a particular limitation to cancer chemotherapy, antibiotic treatment and HIV medication. The ABC (ATP binding cassette) transporters human P-glycoprotein (ABCB1) and the human MRP5 (ABCC5) are involved in multidrug resistance.</p> <p>Results</p> <p>In order to elucidate structural and molecular concepts of multidrug resistance, we have constructed a molecular model of the ATP-bound outward facing conformation of the human multidrug resistance protein ABCB1 using the Sav1866 crystal structure as a template, and compared the ABCB1 model with a previous ABCC5 model. The electrostatic potential surface (EPS) of the ABCB1 substrate translocation chamber, which transports cationic amphiphilic and lipophilic substrates, was neutral with negative and weakly positive areas. In contrast, EPS of the ABCC5 substrate translocation chamber, which transports organic anions, was generally positive. Positive-negative ratios of amino acids in the TMDs of ABCB1 and ABCC5 were also analyzed, and the positive-negative ratio of charged amino acids was higher in the ABCC5 TMDs than in the ABCB1 TMDs. In the ABCB1 model residues Leu65 (transmembrane helix 1 (TMH1)), Ile306 (TMH5), Ile340 (TMH6) and Phe343 (TMH6) may form a binding site, and this is in accordance with previous site directed mutagenesis studies.</p> <p>Conclusion</p> <p>The Sav1866 X-ray structure may serve as a suitable template for the ABCB1 model, as it did with ABCC5. The EPS in the substrate translocation chambers and the positive-negative ratio of charged amino acids were in accordance with the transport of cationic amphiphilic and lipophilic substrates by ABCB1, and the transport of organic anions by ABCC5.</p

Modulation of high affinity ATP-dependent cyclic nucleotide transporters by specific and non-specific cyclic nucleotide phosphodiesterase inhibitors

AbstractIntracellular cyclic nucleotides are eliminated by phosphodiesterases (PDEs) and by ATP Binding cassette transporters such as ABCC4 and ABCC5. PDE5 and ABCC5 have similar affinity for cGMP whereas ABCC5 has much higher affinity for cGMP compared with cAMP. Since the substrate (cGMP) is identical for these two eliminatory processes it is conceivable that various PDE inhibitors also modulate ABCC5-transport. Cyclic GMP is also transported by ABBC4 but the affinity is much lower with a Km 50–100 times higher than for that of ABBCC5. The present study aimed to determine Ki-values for specific or relative specific PDE5 inhibitors (vardenafil, tadalafil, zaprinast and dipyridamole) and the non-specific PDE inhibitors (IBMX, caffeine and theophylline) for ABCC5 andABCC4 transport. The transport of [3H]-cGMP (2µM) was concentration-dependently inhibited with the following Ki-values: vardenafil (0.62µM), tadalafil (14.1µM), zaprinast (0.68µM) and dipyridamole (1.2µM), IBMX (10µM), caffeine (48µM) and theophylline (69µM). The Ki-values for the inhibition of the [3H]-cAMP (2µM) transport were: vardenafil (3.4µM), tadalafil (194µM), zaprinast (2.8µM), dipyridamole (5.5µM), IBMX (16µM), caffeine (41µM) and theophylline (85µM). The specificity for ABCC5 we defined as ratio between Ki-values for inhibition of [3H]-cGMP and [3H]-cAMP transport. Tadalafil showed the highest specificity (Ki-ratio: 0.073) and caffeine the lowest (Ki-ratio: 1.2)

A short update on the structure of drug binding sites on neurotransmitter transporters

<p>Abstract</p> <p>Background</p> <p>The dopamine (DAT), noradrenalin (NET) and serotonin (SERT) transporters are molecular targets for different classes of psychotropic drugs. Cocaine and the SSRI (<it>S</it>)-citalopram block neurotransmitter reuptake competitively, but while cocaine is a non-selective reuptake inhibitor, (<it>S</it>)-citalopram is a selective SERT inhibitor.</p> <p>Findings</p> <p>Here we present comparisons of the binding sites and the electrostatic potential surfaces (EPS) of DAT, NET and SERT homology models based on two different LeuT_Aatemplates; with a substrate (leucine) in an occluded conformation (PDB id <ext-link ext-link-id="2a65" ext-link-type="pdb">2a65</ext-link>), and with an inhibitor (tryptophan) in an open-to-out conformation (PDB id <ext-link ext-link-id="3f3a" ext-link-type="pdb">3f3a</ext-link>). In the occluded homology models, two conserved aromatic amino acids (tyrosine and phenylalanine) formed a gate between the putative binding pockets, and this contact was interrupted in the open to out conformation. The EPS of DAT and NET were generally negative in the vestibular area, whereas the EPS of the vestibular area of SERT was more neutral.</p> <p>Conclusions</p> <p>The findings presented here contribute as an update on the structure of the binding sites of DAT, NET and SERT. The updated models, which have larger ligand binding site areas than models based on other templates, may serve as improved tools for virtual ligand screening.</p

Molecular model of the outward facing state of the human P-glycoprotein (ABCB1), and comparison to a model of the human MRP5 (ABCC5)-3

<p><b>Copyright information:</b></p><p>Taken from "Molecular model of the outward facing state of the human P-glycoprotein (ABCB1), and comparison to a model of the human MRP5 (ABCC5)"</p><p>http://www.tbiomed.com/content/4/1/33</p><p>Theoretical Biology & Medical Modelling 2007;4():33-33.</p><p>Published online 6 Sep 2007</p><p>PMCID:PMC2211457.</p><p></p> Sav1866, while the next branch (the "ABCC-branch") includes ABCC5

Molecular model of the outward facing state of the human P-glycoprotein (ABCB1), and comparison to a model of the human MRP5 (ABCC5)-2

<p><b>Copyright information:</b></p><p>Taken from "Molecular model of the outward facing state of the human P-glycoprotein (ABCB1), and comparison to a model of the human MRP5 (ABCC5)"</p><p>http://www.tbiomed.com/content/4/1/33</p><p>Theoretical Biology & Medical Modelling 2007;4():33-33.</p><p>Published online 6 Sep 2007</p><p>PMCID:PMC2211457.</p><p></p> the intracellular side with blue areas indicating positive areas and red areas indicating negative areas. TMHs are displayed as green ribbons. TMH numbering is indicated in white boxes

Molecular model of the outward facing state of the human P-glycoprotein (ABCB1), and comparison to a model of the human MRP5 (ABCC5)-1

<p><b>Copyright information:</b></p><p>Taken from "Molecular model of the outward facing state of the human P-glycoprotein (ABCB1), and comparison to a model of the human MRP5 (ABCC5)"</p><p>http://www.tbiomed.com/content/4/1/33</p><p>Theoretical Biology & Medical Modelling 2007;4():33-33.</p><p>Published online 6 Sep 2007</p><p>PMCID:PMC2211457.</p><p></p>olor code of the models is blue via white to red from N-terminal to C-terminal