NMR structure note: solution structure of Ca(2+) binding domain 2B of the third isoform of the Na(+)/Ca (2+) exchanger

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Structural and dynamic aspects of ca2+ and mg2+ binding of the regulatory domains of the na+/ca2+ exchanger

Contains fulltext : 94113.pdf (publisher's version ) (Closed access)Intracellular Ca2+ regulates the activity of the NCX (Na+/Ca2+ exchanger) through binding to the cytosolic CBD (Ca2+-binding domain) 1 and CBD2. In vitro studies of the structure and dynamics of CBD1 and CBD2, as well as studies of their kinetics and thermodynamics of Ca2+ binding, greatly enhanced our understanding of NCX regulation. We describe the fold of the CBDs in relation to other known structures and review Ca2+ binding of the different CBD variants from a structural perspective. We also report on new findings concerning Mg2+ binding to the CBDs and finally we discuss recent results on CBD1-CBD2 interdomain interactions

Expression and purification of the c-terminal fragments of trpv5/6 channels

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Structural basis for Ca2+ regulation in the Na+/Ca2+ exchanger

Contains fulltext : 36511.pdf (publisher's version ) (Closed access)Binding of Na+ and Ca2+ ions to the large cytosolic loop of the Na+/Ca2+ exchanger (NCX) regulates its ion transport across the plasma membrane. We determined the solution structures of two Ca2+-binding domains (CBD1 and CBD2) that, together with an alpha-catenin-like domain (CLD) form the regulatory exchanger loop. CBD1 and CBD2 constitute a novel Ca2+-binding motif and are very similar in the Ca2+-bound state. Strikingly, in the absence of Ca2+ the upper half of CBD1 unfolds while CBD2 maintains its structural integrity. Together with a sevenfold higher affinity for Ca2+ this suggests that CBD1 is the primary Ca2+ sensor. Specific point mutations in either domain largely allow the interchange of their functionality and uncover the mechanism underlying Ca2+ sensing in NCX

CEESY: Characterizing the conformation of unobservable protein states

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Role of structural and dynamical plasticity in Sin3: the free PAH2 domain is a folded module in mSin3B

The co-repressor Sin3 is the essential scaffold protein of the Sin3/HDAC co-repressor complex, which is recruited to the DNA by a diverse group of transcriptional repressors, targeting genes involved in the regulation of the cell cycle, proliferation and differentiation. Sin3 contains four repeats commonly denoted as paired amphipathic helix (PAH1-4) domains that provide the principal interaction surface for various repressors. Here, we present the first structure of the free state of the PAH2 domain and discuss its implications for interaction with the repressors. The unbound conformation is very similar to the conformation observed when bound to either the Mad1 or HBP1 repressor, suggesting that the PAH2 domain serves as a template that guides proper folding of the unstructured repressor. The free PAH2 domain shows micro- to millisecond conformational exchange between the folded, major state and a partially unfolded, minor state. Upon complex formation, we observe a significant decrease in fast time-scale flexibility of local regions of the protein, correlated with the formation of intermolecular contacts, and an overall decrease in the slow time-scale conformational exchange. On the basis of our data and using a multiple sequence alignment of all PAH domains, we suggest that the PAH1, PAH2 and PAH3 domains form pre-folded binding modules in full-length Sin3 like beads-on-a-string, and act as folding templates for the interaction domains of their targets

Measurements of structural constraints

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The second ca(2+)-binding domain of ncx1 binds mg(2+) with high affinity

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