Tuning the pH profile of β-glucuronidase by rational site-directed mutagenesis for efficient transformation of glycyrrhizin

Qiaofeng Li; Tian Jiang; Rui Liu; Xudong Feng; Chun Li

doi:10.1007/s00253-019-09790-3

Tuning the pH profile of β-glucuronidase by rational site-directed mutagenesis for efficient transformation of glycyrrhizin

Qiaofeng Li, Tian Jiang, Rui Liu, Xudong Feng^*, Chun Li

^*Corresponding author for this work

School of Chemistry and Chemical Engineering

Research output: Contribution to journal › Article › peer-review

25 Citations (Scopus)

Abstract

In this study, we aimed to shift the optimal pH of acidic β-glucuronidase from Aspergillus oryzae Li-3 (PGUS) to the neutral region by site-directed mutagenesis, thus allowing high efficient biotransformation of glycyrrhizin (GL) into glycyrrhetinic acid (GA) under higher pH where the solubility of GL could be greatly enhanced. Based on PGUS structure analysis, five critical aspartic acid and glutamic acid residues were replaced with arginine on the surface to generate a variant 5Rs with optimal pH shifting from 4.5 to 6.5. The catalytic efficiency (k_cat /K_m) value of 5Rs at pH 6.5 was 10.7-fold higher than that of PGUS wild-type at pH 6.5, even 1.4-fold higher than that of wild-type at pH 4.5. Molecular dynamics simulation was performed to explore the molecular mechanism for the shifted pH profile and enhanced pH stability of 5Rs.

Original language	English
Pages (from-to)	4813-4823
Number of pages	11
Journal	Applied Microbiology and Biotechnology
Volume	103
Issue number	12
DOIs	https://doi.org/10.1007/s00253-019-09790-3
Publication status	Published - 18 Jun 2019

Keywords

Arginine
Rational design
pH shift
pH stability
β-Glucuronidase

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10.1007/s00253-019-09790-3

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title = "Tuning the pH profile of β-glucuronidase by rational site-directed mutagenesis for efficient transformation of glycyrrhizin",

abstract = "In this study, we aimed to shift the optimal pH of acidic β-glucuronidase from Aspergillus oryzae Li-3 (PGUS) to the neutral region by site-directed mutagenesis, thus allowing high efficient biotransformation of glycyrrhizin (GL) into glycyrrhetinic acid (GA) under higher pH where the solubility of GL could be greatly enhanced. Based on PGUS structure analysis, five critical aspartic acid and glutamic acid residues were replaced with arginine on the surface to generate a variant 5Rs with optimal pH shifting from 4.5 to 6.5. The catalytic efficiency (kcat /Km) value of 5Rs at pH 6.5 was 10.7-fold higher than that of PGUS wild-type at pH 6.5, even 1.4-fold higher than that of wild-type at pH 4.5. Molecular dynamics simulation was performed to explore the molecular mechanism for the shifted pH profile and enhanced pH stability of 5Rs.",

keywords = "Arginine, Rational design, pH shift, pH stability, β-Glucuronidase",

author = "Qiaofeng Li and Tian Jiang and Rui Liu and Xudong Feng and Chun Li",

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AU - Li, Qiaofeng

AU - Jiang, Tian

AU - Liu, Rui

AU - Feng, Xudong

AU - Li, Chun

PY - 2019/6/18

Y1 - 2019/6/18

N2 - In this study, we aimed to shift the optimal pH of acidic β-glucuronidase from Aspergillus oryzae Li-3 (PGUS) to the neutral region by site-directed mutagenesis, thus allowing high efficient biotransformation of glycyrrhizin (GL) into glycyrrhetinic acid (GA) under higher pH where the solubility of GL could be greatly enhanced. Based on PGUS structure analysis, five critical aspartic acid and glutamic acid residues were replaced with arginine on the surface to generate a variant 5Rs with optimal pH shifting from 4.5 to 6.5. The catalytic efficiency (kcat /Km) value of 5Rs at pH 6.5 was 10.7-fold higher than that of PGUS wild-type at pH 6.5, even 1.4-fold higher than that of wild-type at pH 4.5. Molecular dynamics simulation was performed to explore the molecular mechanism for the shifted pH profile and enhanced pH stability of 5Rs.

AB - In this study, we aimed to shift the optimal pH of acidic β-glucuronidase from Aspergillus oryzae Li-3 (PGUS) to the neutral region by site-directed mutagenesis, thus allowing high efficient biotransformation of glycyrrhizin (GL) into glycyrrhetinic acid (GA) under higher pH where the solubility of GL could be greatly enhanced. Based on PGUS structure analysis, five critical aspartic acid and glutamic acid residues were replaced with arginine on the surface to generate a variant 5Rs with optimal pH shifting from 4.5 to 6.5. The catalytic efficiency (kcat /Km) value of 5Rs at pH 6.5 was 10.7-fold higher than that of PGUS wild-type at pH 6.5, even 1.4-fold higher than that of wild-type at pH 4.5. Molecular dynamics simulation was performed to explore the molecular mechanism for the shifted pH profile and enhanced pH stability of 5Rs.

KW - Arginine

KW - Rational design

KW - pH shift

KW - pH stability

KW - β-Glucuronidase

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IS - 12

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Tuning the pH profile of β-glucuronidase by rational site-directed mutagenesis for efficient transformation of glycyrrhizin

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Keywords

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