title: Simple discovery of bacterial biocatalysts from environmental samples through functional metaproteomics.
abstract: Bacterial biocatalysts play a key role in our transition to a bio-based, post-petroleum economy. However, the discovery of new biocatalysts is currently limited by our ability to analyze genomic information and our capacity of functionally screening for desired activities. Here, we present a simple workflow that combines functional metaproteomics and metagenomics, which facilitates the unmediated and direct discovery of biocatalysts in environmental samples. To identify the entirety of lipolytic biocatalysts in a soil sample contaminated with used cooking oil, we detected all proteins active against a fluorogenic substrate in sample's metaproteome using a 2D-gel zymogram. Enzymes' primary structures were then deduced by tryptic in-gel digest and mass spectrometry of the active protein spots, searching against a metagenome database created from the same contaminated soil sample. We then expressed one of the novel biocatalysts heterologously in Escherichia coli and obtained proof of lipolytic activity.
authors.affiliation: Ruhr-Universität Bochum, Institute for Biochemistry and Pathobiochemistry - Microbial Biochemistry, Universitätsstr. 150, 44780, Bochum, Germany.
authors.forename: Premankur
authors.initials: P
authors.lastname: Sukul
authors.affiliation: Ruhr-Universität Bochum, Applied Microbiology, Universitätsstr. 150, 44780, Bochum, Germany.
authors.forename: Sina
authors.initials: S
authors.lastname: Schäkermann
authors.affiliation: Ruhr-Universität Bochum, Applied Microbiology, Universitätsstr. 150, 44780, Bochum, Germany.
authors.forename: Julia E
authors.initials: JE
authors.lastname: Bandow
authors.affiliation: Ruhr-Universität Bochum, Institute for Biochemistry and Pathobiochemistry - Microbial Biochemistry, Universitätsstr. 150, 44780, Bochum, Germany.
authors.forename: Anna
authors.initials: A
authors.lastname: Kusnezowa
authors.affiliation: Ruhr-Universität Bochum, Lehrstuhl für Allgemeine und Molekulare Botanik, Universitätsstr. 150, 44780, Bochum, Germany.
authors.forename: Minou
authors.initials: M
authors.lastname: Nowrousian
authors.affiliation: Ruhr-Universität Bochum, Institute for Biochemistry and Pathobiochemistry - Microbial Biochemistry, Universitätsstr. 150, 44780, Bochum, Germany. lars.leichert@ruhr-uni-bochum.de.
authors.forename: Lars I
authors.initials: LI
authors.lastname: Leichert
mesh_terms: D004787:Environmental Pollution; D052918:Environmental Restoration and Remediation / Q000379:methods*; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics / Q000378:metabolism; D008049:Lipase / Q000235:genetics* / Q000378:metabolism; D050356:Lipid Metabolism / Q000235:genetics*; D008055:Lipids / Q000032:analysis; D008066:Lipolysis / Q000502:physiology; D013058:Mass Spectrometry / Q000379:methods; D056186:Metagenomics / Q000379:methods*; D040901:Proteomics / Q000379:methods*; D012988:Soil Microbiology
chemical_list: D008055:Lipids; D008049:Lipase
keywords: Biocatalyst; Lipase; Metagenomics; Metaproteomics; Zymogram