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Structure and characterization of Aspergillus fumigatus lipase B with a unique, oversized regulatory subdomain
Author(s) -
Huang Weiqian,
Lan Dongming,
Popowicz Grzegorz M.,
Zak Krzysztof M.,
Zhao Zexin,
Yuan Hong,
Yang Bo,
Wang Yonghua
Publication year - 2019
Publication title -
the febs journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.981
H-Index - 204
eISSN - 1742-4658
pISSN - 1742-464X
DOI - 10.1111/febs.14814
Subject(s) - lipase , thermostability , candida antarctica , aspergillus fumigatus , chemistry , fungal protein , protein engineering , active site , hydrolase , biochemistry , enzyme , stereochemistry , mutant , biology , gene , microbiology and biotechnology
Fungal lipases are efficient and environment‐friendly biocatalysts for many industrially relevant processes. One of the most widely applied lipases in the manufacturing industry is Candida antarctica lipase B ( CALB ). Here, we report the biochemical and structural characterization of a novel CALB ‐like lipase from an important human pathogen— Aspergillus fumigatus ( AFLB ), which has high sn‐1,3‐specificity toward triolein. AFLB crystal structure displays a CALB ‐like catalytic domain and hosts a unique tightly closed ‘lid’ domain that contains a disulfide bridge, as well as an extra N‐terminal subdomain composed of residues 1–128 (including the helix α1–α5 located above the active site). To gain insight into the function of this novel lid and N‐terminal subdomain, we constructed and characterized a series of mutants in these two domains. Deleting the protruding bulk lid's residues, replacing the bulk and tight lid with a small and loose lid from CALB , or breaking the disulfide bridge increased the affinity of CALB for glyceride substrates and improved its catalytic activity, along with the loss of enzyme fold stability and thermostability. N‐terminal truncation mutants revealed that the N‐terminal peptide (residues 1–59) is a strong inhibitor of AFLB binding to lipid films. This peptide thus limits AFLB 's penetration power and specific activity, revealing a unique enzyme activity regulatory mechanism. Our findings on the functional and structural properties of AFLB provide a better understanding of the functions of the CALB ‐like lipases and pave the way for its future protein engineering. Database Structural data are available in the Protein Data Bank under the accession numbers 6IDY .