Aspergillus niger lipase: Heterologous expression in Pichia pastoris, molecular modeling prediction and the importance of the hinge domains at both sides of the lid domain to interfacial activation.
Aspergillus niger lipase (ANL) is an important biocatalyst in the food processing industry. However, there is no report of its detailed three-dimensional structure because of difficulties in crystallization. In this article, based on experimental data and bioinformational analysis results, the structural features of ANL were simulated. Firstly, two recombinant ANLs expressed in Pichia pastoris were purified to homogeneity and their corresponding secondary structure compositions were determined by circular dichroism spectra. Secondly, the primary structure, the secondary structure and the three-dimensional structure of ANL were modeled by comparison with homologous lipases with known three-dimensional structures using the BioEdit software, lipase engineering database (http://www.led.uni-stuttgart.de/), PSIPRED server and SwissModel server. The predicted molecular structure of ANL presented typical features of the alpha/beta hydrolase fold including positioning of the putative catalytic triad residues and the GXSXG signature motif. Comparison of the predicted three-dimensional structure of ANL with the X-ray three-dimensional structure of A. niger feruloyl esterase showed that the functional difference of interfacial activation between lipase and esterase was concerned with the difference in position of the lid. Our three-dimensional model of ANL helps to modify lipase structure by protein engineering, which will further expand the scope of application of ANL.
Shu
Zhengyu
Z
Engineering Research Center of Industrial Microbiology, Ministry of Education, Fujian Normal University, Fuzhou Fujian 350108, PR China. shuzhengyu@gmail.com
Duan
Mojie
M
Yang
Jiangke
J
Xu
Li
L
Yan
Yunjun
Y
D000595:Amino Acid Sequence; D001234:Aspergillus niger / Q000201:enzymology*; D004789:Enzyme Activation; D005656:Fungal Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015870:Gene Expression; D021621:Imaging, Three-Dimensional; D008049:Lipase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D010843:Pichia / Q000235:genetics* / Q000378:metabolism; D017434:Protein Structure, Tertiary; D016415:Sequence Alignment
D005656:Fungal Proteins; D008049:Lipase