Mouse anti-free-Bata-HCG Monoclonal antibody, clone NM11I15

Given the interaction of the inositol 1,4,5-trisphosphate receptor (IP3R) with chromogranins A (CGA) and B (CGB), two major Ca2+ storage proteins of secretory granules that have been shown to be IP3-sensitive intracellular Ca2+ store of neuroendocrine cells, we have investigated the potential interaction of the intraluminal loop regions of the IP3R with both intact CGB and the conserved near N-terminal region of CGB. The interaction studies carried out with CGB and glutathione S-transferase fusion proteins of intraluminal loop regions of bovine type 1 IP3R showed that CGB interacts with intraluminal loop 3-2 (the second loop formed between transmembrane regions 5 and 6) of the IP3R at both pH 5.5 and 7.5. Analytical ultracentrifugation studies also indicated-that CGB interacts with the same intraluminal loop region of the IP3R and the interaction was much stronger than that between CGA and the loop. Moreover, the conserved near N-terminal region of CGB also interacted with the intraluminal loop region of the IP3R. The CGB interaction with the IP3R intraluminal loop peptide at pH 7.5 showed a Delta G(0) value of -8.1 kcal/ mol at 37 degrees C for a 1:1 stoichiometry, indicating a K-d of similar to 1.9 mu M. These results give insight into the molecular organization of the IP3-sensitive Ca2+ store.

Chromogranins A and B (CGA and CGB) are high capacity, low affinity calcium (Ca2+) storage proteins found in many cell types most often associated with secretory granules of secretory cells but also with the endoplasmic reticulum ( ER) lumen of these cells. Both CGA and CGB associate with inositol 1,4,5-trisphosphate receptor (InsP(3)R) in a pH-dependent manner. At an intraluminal pH of 5.5, as found in secretory vesicles, both CGA and CGB bind to the InsP(3)R. When the intraluminal pH is 7.5, as found in the ER, CGA totally dissociates from InsP(3)R, whereas CGB only partially dissociates. To investigate the functional consequences of the interaction between the InsP(3)R and CGB monomers or CGA/ CGB heteromers, purified mouse InsP(3)R type I were fused to planar lipid bilayers and activated by 2 muM InsP(3). In the presence of luminal CGB monomers or CGA/ CGB heteromers the InsP(3)R/Ca2+ channel open probability and mean open time increased significantly. The channel activity remained elevated when the pH was changed to 7.5, a reflection of CGB binding to the InsP3R even at pH 7.5. These results suggest that CGB may play an important modulatory role in the control of Ca2+ release from the ER. Furthermore, the difference in the ability of CGA and CGB to regulate the InsP(3)R/Ca2+ channel and the variability of CGA/ CGB ratios could influence the pattern of InsP(3)-mediated Ca2+ release.