DIMERIZATION REGULATES THE ENZYMATIC-ACTIVITY OF ESCHERICHIA-COLI OUTER-MEMBRANE PHOSPHOLIPASE-A

The outer membrane phospholipase A (OMPLA) of Escherichia coli is pres ent in a dormant state in the cell envelope, The enzyme is activated b y various processes, which have in common that they perturb the outer membrane. Kinetic experiments, chemical cross-linking, and analytical ultracentrifugation were carried out with purified, detergent-solubili zed OMPLA to understand the underlying mechanism that results in activ ation, Under conditions in which the enzyme displayed full activity, O MPLA was dimeric. High detergent concentrations or very dilute protein concentrations resulted in low specific activity of the enzyme, and u nder those conditions the enzyme was monomeric, The cofactor Ca2+ was required for dimerization, Covalent modification of the active site se rine with hexadecylsulfonylfluoride resulted in stabilization of the d imeric form and a loss of the absolute calcium requirement for dimeriz ation. The results of these experiments provide evidence for dimerizat ion as the molecular mechanism by which the enzymatic activity of OMPL A is regulated, This dimerization probably plays a role in vivo as wel l. Data from chemical cross-linking on whole cells indicate that OMPLA is present in the outer membrane as a monomer and that activation of the enzyme induces dimerization concurrent with the appearance of enzy matic activity.