Oligomerization of the polycystin-2 C-terminal tail and effects on its Ca2+-binding properties.
Bottom Line: Consequently, trimerization does not further improve the affinity of Ca(2+) binding in the SUPC2 Ccore relative to the isolated EF-hand domain.Our study provides a structural basis for understanding the Ca(2+)-dependent regulation of the PC2 channel by its cytosolic C-terminal domain.The improved methodology also serves as a good strategy to characterize other Ca(2+)-binding proteins.
Affiliation: From the Departments of Laboratory Medicine, Pharmacology, and firstname.lastname@example.org.Show MeSH
Related in: MedlinePlus
Mentions: Characterizing the Ca2+-binding elements postulated to reside within the PC2 C-terminal tail is crucial for understanding the mechanism of channel regulation. Based on the bell-shaped response of open probability to increasing Ca2+ concentration, it can be hypothesized that separate Ca2+-binding sites are responsible for different sides of the bell-shaped curve (4, 5, 7, 9, 16). The EF-hand domain (amino acids 719–798 of human PC2) contains one Ca2+-binding site, which is important for the Ca2+-dependent regulation of PC2 channel (7, 9). However, the isolated EF-hand domain binds Ca2+ very weakly, with a KD of 461 μm, outside the physiologic range of cytosolic Ca2+ concentrations (17). Therefore, it was speculated that there could be additional Ca2+-response elements located in the C-terminal domain outside the EF-hand region (7). Moreover, a series of acidic residues located in the loop region (linker 2-L2) (Fig. 1A) connecting the EF-hand and coiled-coil domain share a sequence similar to the Ca2+-bowl structure found in the BK (big potassium) channel (18, 19). It was important to determine the Ca2+-binding profiles of HPC2 Cterm because this would address the question of whether there are additional C-terminal Ca2+-binding sites outside the EF-hand domain and if the Ca2+-binding affinity of HPC2 Cterm is within the physiologic range (nanomolar to low micromolar). However, the Ca2+-binding profile of the HPC2 Cterm was not fully determined in previous studies using isothermal titration calorimetry (ITC) due to the presence of residual Ca2+ and protein aggregates (7).
Affiliation: From the Departments of Laboratory Medicine, Pharmacology, and email@example.com.