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 (K(d)) 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 and 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.