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The Structure and Enzyme Characteristics of a Recombinant Leucine Aminopeptidase rLap1 from Aspergillus sojae and Its Application in Debittering

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Abstract

A leucine aminopeptidase Lap1 was cloned from Aspergillus sojae GIM3.30. The truncated Lap1 without a signal peptide was over-expressed in P. pastoris, and the enzymatic characteristics of recombinant Lap1 (rLap1) were tested. The rLap1 was about 36.7 kDa with an optimal pH 8.0 and optimal temperature 50 °C for substrate Leu-p-nitroanilide and it sustained 50 % activity after 1 h incubation at 50 °C. The activity of rLap1 was significantly inhibited by EDTA, whereas Co2+, Mn2+, and Ca2+ ions, but not Zn2+ ions, restored its activity. rLap1 showed the highest activity against Arg-pNA and then Leu-, Lys-, Met-, and Phe-pNA. The 3D structure of rLap1 showed it had a conserved functional charge/dipole complex and a hydrogen bond network of Zn2-D179-S228-Q177-D229-S158 around its active center. An acidic Asp residue was found at the bottom of the substrate binding pocket, which explains its preference for basic N-terminal amino acid substrates such as Arg and Lys. rLap1 improved the degree of hydrolysis of casein and soy protein hydrolysates and also decreased their bitterness, indicating its potential utility in food production.

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Acknowledgments

This work was financially supported by Guangdong Province of China Science and Technology Projects (Project No. 2012B020311003, No. 2013B010404003, No. 2014A010107005).

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Authors and Affiliations

  1. Guangdong Provincial Key Laboratory of Fermentation and enzyme engineering, School of Bioscience and Bioengineering, South China University of Technology, Guangzhou Higher Education Mega Center, Panyu District, Guangzhou, Guangdong, People’s Republic of China, 510006

    Wei-Qian Huang, Li-Fen Zhong, Zhi-Zhong Meng, Zi-Juan You & Xiao-Chun Luo

  2. Department of Food and Bioengineering, Guangdong Industry Technical College, Guangzhou, Guangdong, People’s Republic of China, 510300

    Jia-Zhou Li

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  1. Wei-Qian Huang
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Huang, WQ., Zhong, LF., Meng, ZZ. et al. The Structure and Enzyme Characteristics of a Recombinant Leucine Aminopeptidase rLap1 from Aspergillus sojae and Its Application in Debittering. Appl Biochem Biotechnol 177, 190–206 (2015). https://doi.org/10.1007/s12010-015-1737-5

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  • DOI: https://doi.org/10.1007/s12010-015-1737-5

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