Fig. 6

Schematic guideline for the activity assay. Based on all data collected, four major factors were identified to adjust the assay: ionic strength of the buffer, buffer ion denticity, buffer pH and substrate concentration. The green and orange areas correspond to the region lowest and highest LPMO activity, respectively. Black arrows indicate the increase or decrease in activity by adjusting a factor. As a good starting point for the assay, we recommend 100 mM sodium acetate, pH 6.0, 500 µM hydrocoerulignone as chromogenic substrate and 100 µM hydrogen peroxide as cosubstrate. The area around the recommended conditions indicates conditions in which LPMO activity can be detected, but is not optimized for maximum reliability. This region can be used to characterize the LPMO’s behavior under different conditions, e.g., for a pH profile. The area around the recommended conditions indicates conditions in which LPMO activity can be detected, but is not optimized for maximum reliability. This region can be used to characterize the LPMO’s behavior under different conditions, e.g., for a pH profile. We suggest to start with small changes to our recommended conditions to not end up in the green or orange region, where either no activity can be detected or the activity is too high and autoxidation inactivates LPMO too fast