High capacity, low affinity Ca²⁺ binding of chromogranin A: Relationship between the pH-induced conformational change and Ca²⁺ binding property

Chromogranin A, the major intravesicular protein of adrenal chromaffin granules, bound Ca²⁺ in a pH-dependent manner. Both the maximal binding and affinity of chromogranin A for Ca²⁺ were dependent on pH. Chromogranin A bound 670 nmol of Ca²⁺/mg (32 mol/ mol) and 1150 nmol of Ca²⁺/mg (55 mol/mol) at pH 7.5 and 5.5, respectively, with dissociation constants (Kd) of 2.7 and 4 mM. This pH dependence probably reflects different conformations of the protein at the two pH values. Conformational differences of chromogranin A at two different pH values were demonstrated by limited tryptic digestion patterns confirming previous results obtained by circular dichroism spectroscopy (Yoo, S. H., and Albanesi, J. P. (1990) J. Biol. Chem. 265, 14414-14421). Sedimentation equilibrium studies revealed the native molecular mass of chromogranin A to be 100 kDa at pH 7.5 ad 192 kDa at pH 5.5, indicating dimeric and tetrameric states of the protein at the two pH levels. We postulate that the pH- and Ca²⁺-induced conformational changes of chromogranin A may have a role both in the regulation of Ca²⁺ release of chromaffin granules and in the early stages of secretory vesicle biogenesis.