A Novel CalB-Type Lipase Discovered by Fungal Genomes Mining

Vaquero, María Eugenia; Eugenio, Laura I. de; Martı́nez, Marı́a Jesús; Barriuso, Jorge

doi:10.1371/journal.pone.0124882

Cited by 12 publications

(8 citation statements)

References 29 publications

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“…D), with highest activity values toward tributyrin (TC4), followed by trioctanoin (TC8), laurin (TC12), triacetin (TC2), and olive oil. Results show that AFLB exhibited higher activity toward short‐chain triglycerides, similar to that reported CALB‐like lipase CALB, Uml2, and PlicB . Most well‐characterized lipases are either sn ‐1,3 regiospecific or nonregiospecific .…”

Section: Resultssupporting

confidence: 80%

Structure and characterization of Aspergillus fumigatus lipase B with a unique, oversized regulatory subdomain

et al. 2019

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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 http://www.rcsb.org/pdb/search/structidSearch.do?structureId=6IDY.

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Section: Resultssupporting

confidence: 80%

Structure and characterization of Aspergillus fumigatus lipase B with a unique, oversized regulatory subdomain

et al. 2019

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“…With the rapid development of genome sequencing technology, a large number of microbial genes have been analyzed, making it possible to obtain the desired target genes in situ from gene databases such as NCBI, UniProt, and GenBank. In situ enzyme mining can be roughly divided into three steps: (1) An initial template sequence or conservative motif is used as a probe to obtain potential sequences from databases such as NCBI, , UniProt or metagenomics libraries, and then, the sequences are filtered according to homology or whether they contained conserved motifs. (2) Bioinformatics methods are used to continue to analyze potential sequences and narrow down candidate sequences, such as phylogenetic analysis, homology modeling and molecular docking technology.…”

Section: Methodology Workflow For the Cell-free Biosynthesis Systemmentioning

confidence: 99%

Cell-Free Biosynthesis System: Methodology and Perspective of in Vitro Efficient Platform for Pyruvate Biosynthesis and Transformation

Tang

Liao

et al. 2021

ACS Synth. Biol.

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The modification of intracellular metabolic pathways by metabolic engineering has generated many engineered strains with relatively high yields of various target products in the past few decades. However, the unpredictable accumulation of toxic products, the cell membrane barrier, and competition between the carbon flux of cell growth and product synthesis have severely retarded progress toward the industrial-scale production of many essential chemicals. On the basis of an in-depth understanding of intracellular metabolic pathways, scientists intend to explore more sustainable methods and construct a cell-free biosynthesis system in vitro. In this review, the synthesis and application of pyruvate as a platform compound is used as an example to introduce cell-free biosynthesis systems. We systematically summarize a proposed methodology workflow of cell-free biosynthesis systems, including pathway design, enzyme mining, enzyme modification, multienzyme assembly, and pathway optimization. Some new methods, such as machine learning, are also mentioned in this review.

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“…Because both enzymes have similar substrate specificity, they can be expressed from the same microorganism. Majority of lipases and esterases share the same α/β hydrolase fold and are sometimes difficult to differentiate by primary sequence comparisons, structural features, and kinetic parameters (Bracco et al., 2020; Vaquero, de Eugenio, Martínez, & Barriuso, 2015). For instance, Candida antarctica lipase B (Cal B) acts as a lipase when adsorbed to an acylglyceride interface and as an esterase when exposed to an aqueous environment (Benson & Pleiss, 2017).…”

Section: Categories and Applications Of Food Enzymes From Extremophilesmentioning

confidence: 99%

“…Although some lipases display esterase activities, there are some differences between these two classes of enzymes. (Bracco et al, 2020;Vaquero, de Eugenio, Martínez, & Barriuso, 2015). For instance, Candida antarctica lipase B (Cal B) acts as a lipase when adsorbed to an acylglyceride interface and as TA B L E 3 Some esterase producing extremophiles an esterase when exposed to an aqueous environment (Benson & Pleiss, 2017).…”

Section: Esterasesmentioning

confidence: 99%