Mutation and expression of N233C-D506C of cellulase Cel6B from Thermobifida fusca in Escherichia coli

A versatile system for expression and purification of cellulase Cel6B from Thermobifida fusca in Escherichia coli was developed. A double mutation, N233C-D506C, of Cel6B was created to try to increase its thermostability by forming an extra-disulfide bond joining two loops of Cel6B that cover the active site cleft. The predicted mutant enzyme was slightly more thermostable with no CD spectroscopic changes, but a disulfide bond did form and there was a significant decrease in enzyme production.