Addition of a disulfide bond between the N and C termini of B. circulans xylanase stabilizes the protein. The activity at room temperature is doubled and it is largely protected against inactivation at high temperature
Engineering a calcium-independent subtilisn. Native enzyme loses its activity when the calcium-binding loop is deleted. After random mutagenesis, several mutants with low activity are isolated. These are combined into one mutant construct with high activity.
Altering multiple enzyme properties at once. Start with multiple copies of a Bacillus subtilisn gene. Error-prone PCR and then DNA shuffling. Test for high activity at room temp and then stability at high temp