Barttin binds to the outer lateral surface of the ClC-K2 chloride channel
| Autor: | Masato Tajima, Tatemitsu Rai, Sei Sasaki, Shinichi Uchida, Atsushi Hayama |
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| Rok vydání: | 2007 |
| Předmět: |
Models
Molecular Immunoprecipitation Anion Transport Proteins Biophysics Plasma protein binding Antiparallel (biochemistry) Biochemistry Chloride Channels Chlorocebus aethiops Animals Computer Simulation Binding site Molecular Biology COS cells Binding Sites urogenital system Chemistry Cell Membrane Membrane Proteins Cell Biology Transepithelial chloride transport Membrane Models Chemical COS Cells Chloride channel Protein Binding |
| Zdroj: | Biochemical and biophysical research communications. 362(4) |
| ISSN: | 0006-291X |
| Popis: | ClC-K chloride channels belong to the CLC chloride channel family and play an important role in transepithelial chloride transport in the kidney. To be functional, ClC-K channels need to be translocated to the plasma membranes after synthesis; the translocation requires the binding to its beta-subunit, barttin. The binding interaction between barttin and ClC-K channels has not been characterized, although the crystal structure of CLC was resolved. In the present study, we sought to clarify the binding sites of barttin in ClC-K2 by co-immunoprecipitation and immunofluorescence microscopy using various ClC-K2 mutants. The deletion of the carboxy-terminal portion of ClC-K2 up to leucine 91, a construct which contains the B domain alone, showed the binding ability to barttin. Since the CLC channel forms an internal antiparallel structure, domain J corresponds to domain B in the carboxy-terminal half of ClC-K. Accordingly, we made the carboxy-terminal half of ClC-K2 containing domain J and thereafter and its deletion mutants, and performed a similar co-immunoprecipitation study. As a result, only domain J was enough for binding to barttin. Immunofluorescence microscopy confirmed that the domains B and J as well as the full length ClC-K2 could be localized to the plasma membranes only when co-expressed with barttin. These results showed that barttin was able to bind to the domains that constitute the outer lateral surfaces of ClC-K2. This information regarding the binding sites will be useful for designing a new class of diuretics or anti-hypertensive agents that inhibit the interaction of ClC-K and barttin. |
| Databáze: | OpenAIRE |
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