Molecular modeling of p-chlorophenoxyacetic acid binding to the CLC-0 channel.
Molecular simulation techniques were applied to predict the interaction of the CLC-0 Cl(-) channel and the channel-blocking molecule p-chlorophenoxyacetic acid (CPA). A three-dimensional model of the CLC-0 channel was constructed on the basis of the homology with the bacterial Cl(-) channel StCLC, the structure of which has been solved by X-ray crystallography. Docking of the CPA molecule was obtained by using a geometric recognition algorithm, yielding 5000 possible conformations. By restraining the simulation to those conformations in which CPA is near the intracellular mouth of the channel, the CPA-protein complex models were reduced to three sets of conformations, which are interconvertible within 2 ns when molecular dynamics is applied to the system. Point mutations of CLC-0 at three different positions predicted to interact with CPA in these configurations did, however, not greatly alter CPA inhibition, suggesting a deeper final binding location. In the model, binding of CPA to a more internal position in the ionic pathway was obtained by applying a constant force vector to CPA, pushing it toward the center of the channel. This technique allowed us to outline the possible intrachannel pathway of CPA and to describe qualitatively the binding sites and energy barriers of this pathway. The consistency of the obtained models and the experimental data indicates that the CLC-0-CPA complex model is reasonable and can be used in further biological studies, such as rational design of blocking agents of and mutagenesis of CLC Cl(-) channels.
Moran
Oscar
O
Istituto di Biofisica, Sezione di Genova, CNR, Via De Marini, 6, I-16149 Genova, Italy. moran@ge.cnr.it
Traverso
Sonia
S
Elia
Laura
L
Pusch
Michael
M
D015084:2,4-Dichlorophenoxyacetic Acid / Q000031:analogs & derivatives* / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D018118:Chloride Channels / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D018360:Crystallography, X-Ray; D005260:Female; D018698:Glutamic Acid / Q000737:chemistry; D015640:Ion Channel Gating; D008956:Models, Chemical; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009154:Mutation / Q000235:genetics; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D011485:Protein Binding; D011487:Protein Conformation; D012486:Salmonella typhimurium / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D014101:Torpedo / Q000378:metabolism; D014982:Xenopus laevis / Q000378:metabolism
C120501:CLC-0 chloride channel protein, Torpedo californica; D018118:Chloride Channels; D015084:2,4-Dichlorophenoxyacetic Acid; D018698:Glutamic Acid; C007017:4-chlorophenoxyacetic acid