title: A novel Vibrio beta-glucosidase (LamN) that hydrolyzes the algal storage polysaccharide laminarin.
abstract: The metabolic versatility, tractability and rapid growth potential of the Vibrio spp. have made them increasingly attractive systems for investigating carbon cycling in the marine environment. In this study, an in silico subtractive proteomic strategy was used to identify a novel 101 kDa GH3 family β-glucosidase (LamN) that was found in bioluminescent Vibrio campbellii strains capable of utilizing the algal storage glucan laminarin. A heterologous overexpression system verified the sequence-predicted function of LamN as it enabled the growth of Escherichia coli on laminarin as a sole carbon source. Quantitative reverse transcription PCR analyses revealed that V. campbellii grown on laminarin demonstrated a 4- to 314-fold induction of lamN gene expression when compared to the same strains grown on glucose or glycerol. Corresponding tandem mass spectrometric analyses detected LamN protein expression only in cells grown on laminarin. Heterologous expression, purification and biochemical characterization identified LamN as a heat stable laminarinase with β-1,3, β-1,4 and β-1,6 glucosidase activity. Collectively, these data identify an enzyme that may allow V. campbellii to exploit some of the most abundant polysaccharides associated with deteriorating phytoplankton blooms and provide support for the potential involvement of V. campbellii in the formation of bioluminescent milky seas.
authors.affiliation: Center for Bio/Molecular Science and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA.
authors.forename: Zheng
authors.initials: Z
authors.lastname: Wang
authors.affiliation: Center for Bio/Molecular Science and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA.
authors.forename: Kelly L
authors.initials: KL
authors.lastname: Robertson
authors.affiliation: University of Maryland, College Park, MD 20742, USA.
authors.forename: Charles
authors.initials: C
authors.lastname: Liu
authors.affiliation: Center for Bio/Molecular Science and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA.
authors.forename: Jinny L
authors.initials: JL
authors.lastname: Liu
authors.affiliation: Center for Bio/Molecular Science and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA.
authors.forename: Brandy J
authors.initials: BJ
authors.lastname: Johnson
authors.affiliation: Center for Bio/Molecular Science and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA.
authors.forename: Dagmar H
authors.initials: DH
authors.lastname: Leary
authors.affiliation: Nova Research Inc., Alexandria, VA 22308, USA.
authors.forename: Jaimee R
authors.initials: JR
authors.lastname: Compton
authors.affiliation: Department of Microbiology, Prince of Songkla University, Hat Yai 90110, Thailand.
authors.forename: Varaporn
authors.initials: V
authors.lastname: Vuddhakul
authors.affiliation: Center for Bio/Molecular Science and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA.
authors.forename: Patricia M
authors.initials: PM
authors.lastname: Legler
authors.affiliation: Center for Bio/Molecular Science and Engineering, U.S. Naval Research Laboratory, Washington, DC 20375, USA gary.vora@nrl.navy.mil.
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Vora
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000254:growth & development / Q000378:metabolism; D015870:Gene Expression; D005936:Glucans / Q000378:metabolism*; D005947:Glucose / Q000378:metabolism; D005990:Glycerol / Q000378:metabolism; D006868:Hydrolysis; D049449:Luminescence; D010839:Phytoplankton / Q000201:enzymology* / Q000235:genetics; D040901:Proteomics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014733:Vibrio / Q000201:enzymology* / Q000235:genetics; D001617:beta-Glucosidase / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D005936:Glucans; C008247:laminaran; D001617:beta-Glucosidase; D005947:Glucose; D005990:Glycerol
keywords: Phaeocystis; Vibrio campbellii; Vibrio harveyi; bioluminescent milky seas; cellulase; chrysolaminarin; laminarinase