Zymogen‐Enzyme Transformations

Abstract

Transformation of prophospholipase A into phospholipase A is triggered by the tryptic hydrolysis of the Arg 7 ‐Ala 8 bond of the zymogen. This bond is a better substrate for trypsin than the corresponding strategic bonds of other zymogens such as trypsinogen and chymotrypsinogen A or B: K m = 2.2 mM, k cat = 7 s −1 at pH 8, 1°C. Calcium ions which have an essential role in phospholipase catalysis play no role in prophospholipase activation. A comparative physico‐chemical analysis of the zymogen and of the enzyme shows that activation produces only very limited changes in the overall folding of prophospholipase. A structural rearrangement occurs at the N‐terminal end of the precursor. An excellent fluorescence reporter group to follow this conformational modification is the side‐chain of tryptophan‐3 (in phospholipase sequence) which passes from a polar to an apolar environment during activation. The α‐amino group of alanine‐1 (in the phospholipase sequence) which appears on activation is essential for enzyme activity. Selective chemical modification of this function by a number of reagents abolishes phospholipase activity. By analogy with the chymotrypsinogen‐chymotrypsin and the proelastase‐elastase transformations, it is proposed that the α‐amino group is involved in the formation of a salt‐bridge which stabilizes the adequate geometry of the active site. Evidences in favor of this hypothesis include the high pK and the low chemical reactivity of the α‐amino group, together with the resistance of native phospholipase to aminopeptidase degradation.