Enhanced yeast surface display of Β-glucuronidase using dual anchor motifs for high-temperature glycyrrhizin hydrolysis

β-Glucuronidases (GUSs) have been widely used for the modification of natural glucuronide compounds, but the low reusability and stability of GUSs have limited their large-scale use. In this study, PGUS (GUS from Aspergillus oryzae) and AtGUS (GUS from Aspergillus terreus) were independently displayed on the surface of Pichia pastoris via the Pir1 or Agα1 anchoring motif for the efficient preparation of glycyrrhetinic acid (GA) using glycyrrhizin (GL) hydrolysis. The PGUS displayed using Pir1 had 35-fold increased activity than free PGUS. The displayed PGUS also exhibited 6.92-fold higher thermostability at 70°C than free PGUS and retained 75% activity after 10 consecutive runs. Furthermore, the activity of displayed PGUS was increased by 1.44-fold by using the dual complementary anchoring motifs Agα1 and Pir1. A new GL biotransformation process at elevated temperature (60°C) was designed with the displayed PGUS that shows considerable potential for use in the industrial production of GA.