Effects of pH and Ca2+ on heterodimer and heterotetramer formation by chromogranin A and chromogranin B.

The two major proteins of the secretory vesicles of neuroendocrine cells, chromogranin A (CGA) and chromogranin B (CGB), have been shown to undergo pH- and Ca2+-dependent conformational changes and aggregation and have been suggested to play essential roles during secretory vesicle biogenesis in the trans-Golgi network. CGA has been shown to exist primarily in a tetrameric state at pH 5.5 and primarily in a dimeric state at pH 7.5, and CGB has been shown to exist in a monomeric state at both pH 5.5 and pH 7.5. Using purified CGA and CGB, it recently has been shown that CGA interacts with CGB at pH 5.5 (Yoo, S. H.(1996) J. Biol. Chem. 271, 1558-1565). In expanding this investigation, we have studied the temperature dependence of the pH-dependent interaction of CGA and CGB by analytical ultracentrifugation and found that two molecules of CGA bound to two molecules of CGB at pH 5.5 with DeltaG0 values of -43.6 kcal/mol in the absence of Ca2+ at 37 degrees C and -40.3 kcal/mol in the presence of 0.1 mM Ca2+. However, one molecule of CGA bound to one molecule of CGB at pH 7.5 with DeltaG0 values of -13.6 kcal/mol in the absence of Ca2+ at 37 degrees C. The magnitude of DeltaG0 values increased with increasing temperatures at both pH values. However, the values for enthalpy and entropy changes decreased with increasing temperatures in both pH levels, suggesting formation of more ordered structures. In the absence of Ca2+ at pH 5. 5, the heterotetramerization reaction at 37 degrees C was entropically driven, whereas in the presence of Ca2+ (0.1 mM) the heterotetramerization was virtually an enthalpic reaction. On the other hand, the heterodimer formation in the absence of Ca2+ at pH 7. 5 showed large negative enthalpy and entropy changes at 37 degrees C, indicating an enthalpic interaction compensated by entropic changes. In view of the interaction of tetrameric CGA with tetrameric inositol 1,4,5-trisphosphate (IP3) receptor and the existence of heterotetrameric IP3 receptor in the cell, the heterotetramer formation by CGA and CGB not only raises the possibility of interaction between the heterotetrameric chromogranin and heterotetrameric IP3 receptor but also appears to reflect their important roles in the cell.