Gene Expression, Biochemical Characterization of a sn-1, 3 Extracellular Lipase From Aspergillus niger GZUF36 and Its Model-Structure Analysis

Abstract

In this study, a sn-1, 3 extracellular lipases from Aspergillus niger GZUF36 (PEXANL1) was expressed in Pichia pastoris, characterized, and the predicted structural model was analyzed. The optimized culture conditions of P. pastoris showed that the highest lipase activity of 66.5 ± 1.4 U/mL (P < 0.05) could be attained with 1% methanol and 96 h induction time. The purified PEXANL1 exhibited the highest activity at pH 4.0 and 40°C temperature, and its original activity remained unaltered in the majority of the organic solvents (20% v/v concentration). Triton X-100, Tween 20, Tween 80, and SDS at a concentration of 0.01% (w/v) enhanced, and all the metal ions tested inhibited activity of purified PEXANL. The results of ultrasound-assisted PEXANL1 catalyzed synthesis of 1,3-diaglycerides showed that the content of 1,3-diglycerides was rapidly increased to 36.90% with 25 min of ultrasound duration (P < 0.05) and later decreased to 19.93% with 35 min of ultrasound duration. The modeled structure of PEXANL1 by comparative modeling showed α/β hydrolase fold. Structural superposition and molecular docking results validated that Ser162, His274, and Asp217 residues of PEXANL1 were involved in the catalysis. Small-angle X-ray scattering analysis indicated the monomer properties of PEXANL1 in solution. The ab initio model of PEXANL1 overlapped with its modeling structure. This work presents a reliable structural model of A. niger lipase based on homology modeling and small-angle X-ray scattering. Besides, the data from this study will benefit the rational design of suitable crystalline lipase variants in the future.

Characterization of a recombinant Aspergillus niger GZUF36 lipase immobilized by ionic liquid modification strategy

“…The engineering strain P. pastoris GS115 producing the recombinant lipase from A. niger GZUF36 was prepared in our previous work (Xing et al 2021 ) and stored in our laboratory. Fe 3 O 4 , tetraethyl orthosilicate (TEOS), 3-chloropropyltrimethoxysilane (CP-TMS), N-methylimidazole (MI), and p -nitrophenyl palmitate ( p -NPP) were purchased from Shanghai Macklin Biochemical Technology Co., Ltd.…”

“…The production and purification of PEXANL1 were carried out following the methods outlined in our previous study (Xing et al 2021 ). The purified PEXANL1 was stored in a 20 mM Tris buffer (pH 8.0) and used for all immobilization studies.…”

“…Lipases (EC 3.1.1.3) are a vital hydrolase that catalyzes the hydrolysis of the natural substrate lipids at the oil–water interface to produce fatty acids, glycerol, and mono- or diesters. Additionally, lipases are involved in various other reactions, including lipid synthesis, ester exchange, acidolysis, esterolysis, and enantiomeric splitting (Cai et al 2021 ; Xing et al 2021 ; Zhu et al 2021 ). Therefore, lipases are widely used in industries such as surfactants, biodiesel, fine chemicals, wastewater treatment, pharmaceuticals, cosmetics, and flavored foods (Chen et al 2022 ; Salgado et al 2022 ).…”

Fungal lipases as biocatalysts: A promising platform in several industrial biotechnology applications

“…This might be due to the fact that the A. japonicus and Candida rugosa lipases require calcium as a cofactor for their biological activities. However, Xing et al (2021) prove that the activity of lipase from Aspergillus niger GZUF36 is inhibited up to 75% by 5 mM of Ca 2+ . In addition, neither Ca 2+ (5, 10, and 30 mM), Mg 2+ (10, 50, and 100 mM), Na + (0.1, 0.5, and 1 M), nor K + (0.1, 0.5, and 1 M) has an effect on the action of a lipase screened from C. tenuissimum (Saranya & Ramachandra, 2020).…”

“…It maintains 88.86% of its activity in dimethyl sulfoxide (polar solvent). However, this fungal lipase is completely inactivated in dichloromethane (non‐polar solvent) (Xing et al, 2021).…”