Changes in activity of porcine phospholipase A2 brought about by charge engineering of a major structural element to alter stability

We have modified the stability of porcine phospholipase A₂ bycharge engineering. The mutations are situated at the N-terminalof a major helix and are N89D and N89D/E92Q. This engineeringhas significantly altered the activity of the enzyme to aggregatedand monomeric substrates. A N89D/E92K mutant is more stablebut considerably less active than wild type. An N89D mutantis more stable and of similar activity to wild type. The substantialchange in activity may be due to direct interaction of residue92 with aggregated substrate or may be via second calcium binding.Second calcium binding may be more probable as activity againstmonomers is also affected. Additional calcium binding may thereforebe an important way of manipulating the activity of phospholipaseA₂.