title: Genomic and exoproteomic analyses of cold- and alkaline-adapted bacteria reveal an abundance of secreted subtilisin-like proteases.
abstract: Proteases active at low temperature or high pH are used in many commercial applications, including the detergent, food and feed industries, and bacteria specifically adapted to these conditions are a potential source of novel proteases. Environments combining these two extremes are very rare, but offer the promise of proteases ideally suited to work at both high pH and low temperature. In this report, bacteria from two cold and alkaline environments, the ikaite columns in Greenland and alkaline ponds in the McMurdo Dry Valley region, Antarctica, were screened for extracellular protease activity. Two isolates, Arsukibacterium ikkense from Greenland and a related strain, Arsukibacterium sp. MJ3, from Antarctica, were further characterized with respect to protease production. Genome sequencing identified a range of potential extracellular proteases including a number of putative secreted subtilisins. An extensive liquid chromatography-tandem mass spectrometry analysis of proteins secreted by A. ikkense identified six subtilisin-like proteases as abundant components of the exoproteome in addition to other peptidases potentially involved in complete degradation of extracellular protein. Screening of Arsukibacterium genome libraries in Escherichia coli identified two orthologous secreted subtilisins active at pH 10 and 20 °C, which were also present in the A. ikkense exoproteome. Recombinant production of both proteases confirmed the observed activity.
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, Frederiksberg, 1871, Denmark.
authors.forename: Jeanette E
authors.initials: JE
authors.lastname: Lylloff
authors.affiliation: Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Lea B S
authors.initials: LB
authors.lastname: Hansen
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, Frederiksberg, 1871, Denmark.
authors.forename: Morten
authors.initials: M
authors.lastname: Jepsen
authors.affiliation: Interdisciplinary Nanoscience Center and Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Kristian W
authors.initials: KW
authors.lastname: Sanggaard
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, Frederiksberg, 1871, Denmark.
authors.forename: Jan K
authors.initials: JK
authors.lastname: Vester
authors.affiliation: Interdisciplinary Nanoscience Center and Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark.
authors.forename: Jan J
authors.initials: JJ
authors.lastname: Enghild
authors.affiliation: Department of Biology, University of Copenhagen, Copenhagen, Denmark.
authors.forename: Søren J
authors.initials: SJ
authors.lastname: Sørensen
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, Frederiksberg, 1871, Denmark.
authors.forename: Peter
authors.initials: P
authors.lastname: Stougaard
authors.affiliation: Department of Plant and Environmental Sciences, University of Copenhagen, Thorvaldsensvej 40, Frederiksberg, 1871, Denmark.
authors.forename: Mikkel A
authors.identifier: http://orcid.org/0000-0002-8003-4635
authors.initials: MA
authors.lastname: Glaring
mesh_terms: D000468:Alkalies / Q000378:metabolism*; D000864:Antarctic Regions; D002841:Chromatiaceae / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification*; D002853:Chromatography, Liquid; D003080:Cold Temperature; D019295:Computational Biology; D004783:Environmental Microbiology; D004926:Escherichia coli; D023281:Genomics; D006116:Greenland; D040901:Proteomics; D017422:Sequence Analysis, DNA; D013381:Subtilisins / Q000235:genetics / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D000468:Alkalies; D013381:Subtilisins
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