title: Characterization of a sialate-O-acetylesterase (NanS) from the oral pathogen Tannerella forsythia that enhances sialic acid release by NanH, its cognate sialidase.
abstract: Tannerella forsythia, a Gram-negative member of the Bacteroidetes has evolved to harvest and utilize sialic acid. The most common sialic acid in humans is a mono-N-acetylated version termed Neu5Ac (5-N-acetyl-neuraminic acid). Many bacteria are known to access sialic acid using sialidase enzymes. However, in humans a high proportion of sialic acid contains a second acetyl group attached via an O-group, i.e. chiefly O-acetylated Neu5,9Ac2 or Neu5,4Ac2. This diacetylated sialic acid is not cleaved efficiently by many sialidases and in order to access diacetylated sialic acid, some organisms produce sialate-O-acetylesterases that catalyse the removal of the second acetyl group. In the present study, we performed bioinformatic and biochemical characterization of a putative sialate-O-acetylesterase from T. forsythia (NanS), which contains two putative SGNH-hydrolase domains related to sialate-O-acetylesterases from a range of organisms. Purification of recombinant NanS revealed an esterase that has activity against Neu5,9Ac2 and its glycolyl form Neu5Gc,9Ac. Importantly, the enzyme did not remove acetyl groups positioned at the 4-O position (Neu5,4Ac2). In addition NanS can act upon complex N-glycans released from a glycoprotein [erythropoietin (EPO)], bovine submaxillary mucin and oral epithelial cell-bound glycans. When incubated with its cognate sialidase, NanS increased sialic acid release from mucin and oral epithelial cell surfaces, implying that this esterase improves sialic acid harvesting for this pathogen and potentially other members of the oral microbiome. In summary, we have characterized a novel sialate-O-acetylesterase that contributes to the sialobiology of this important human pathogen and has potential applications in the analysis of sialic acid diacetylation of biologics in the pharmaceutical industry.
authors.affiliation: Integrated BioSciences, School of Clinical Dentistry, University of Sheffield, Sheffield, S10 2TA, U.K.
authors.forename: Chatchawal
authors.initials: C
authors.lastname: Phansopa
authors.affiliation: Ludger Ltd, Culham Science Centre, Oxfordshire OX14 3EB, U.K.
authors.forename: Radoslaw P
authors.initials: RP
authors.lastname: Kozak
authors.affiliation: Ludger Ltd, Culham Science Centre, Oxfordshire OX14 3EB, U.K.
authors.forename: Li Phing
authors.initials: LP
authors.lastname: Liew
authors.affiliation: Integrated BioSciences, School of Clinical Dentistry, University of Sheffield, Sheffield, S10 2TA, U.K.
authors.forename: Andrew M
authors.initials: AM
authors.lastname: Frey
authors.affiliation: Integrated BioSciences, School of Clinical Dentistry, University of Sheffield, Sheffield, S10 2TA, U.K.
authors.forename: Thomas
authors.initials: T
authors.lastname: Farmilo
authors.affiliation: Integrated BioSciences, School of Clinical Dentistry, University of Sheffield, Sheffield, S10 2TA, U.K.
authors.forename: Jennifer L
authors.initials: JL
authors.lastname: Parker
authors.affiliation: Department of Molecular Biology and Biotechnology, University of Sheffield, Sheffield S10 2TN, U.K.
authors.forename: David J
authors.initials: DJ
authors.lastname: Kelly
authors.affiliation: Ludger Ltd, Culham Science Centre, Oxfordshire OX14 3EB, U.K.
authors.forename: Robert J
authors.initials: RJ
authors.lastname: Emery
authors.affiliation: Reading School of Pharmacy, University of Reading, Whiteknights, Reading RG6 6AP, U.K.
authors.forename: Rebecca I
authors.initials: RI
authors.lastname: Thomson
authors.affiliation: Ludger Ltd, Culham Science Centre, Oxfordshire OX14 3EB, U.K.
authors.forename: Louise
authors.initials: L
authors.lastname: Royle
authors.affiliation: Ludger Ltd, Culham Science Centre, Oxfordshire OX14 3EB, U.K.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Gardner
authors.affiliation: Ludger Ltd, Culham Science Centre, Oxfordshire OX14 3EB, U.K.
authors.forename: Daniel I R
authors.initials: DI
authors.lastname: Spencer
authors.affiliation: Integrated BioSciences, School of Clinical Dentistry, University of Sheffield, Sheffield, S10 2TA, U.K. g.stafford@sheffield.ac.uk.
authors.forename: Graham P
authors.initials: GP
authors.lastname: Stafford
mesh_terms: D000107:Acetylation; D000115:Acetylesterase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001439:Bacteroides / Q000201:enzymology*; D020134:Catalytic Domain; D002417:Cattle; D045744:Cell Line, Tumor; D004921:Erythropoietin / Q000235:genetics / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D009061:Mouth Mucosa / Q000166:cytology / Q000378:metabolism* / Q000382:microbiology; D009438:Neuraminic Acids / Q000378:metabolism*; D009439:Neuraminidase / Q000737:chemistry / Q000378:metabolism*; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D012794:Sialic Acids / Q000378:metabolism*; D012795:Sialoglycoproteins / Q000737:chemistry / Q000378:metabolism; D052116:Sialomucins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; C554270:EPO protein, human; D009438:Neuraminic Acids; D011134:Polysaccharides; D011994:Recombinant Proteins; D012794:Sialic Acids; D012795:Sialoglycoproteins; D052116:Sialomucins; D004921:Erythropoietin; D000115:Acetylesterase; C046298:sialate O-acetylesterase; D009439:Neuraminidase
keywords: Bacteroidetes; acetlyesterase; carbohydrate-active enzyme; glycans; oral cavity; sialic acid