title: Glycan specificity of neuraminidases determined in microarray format.
abstract: Neuraminidases hydrolytically remove sialic acids from glycoconjugates. Neuraminidases are produced by both humans and their pathogens, and function in normal physiology and in pathological events. Identification of neuraminidase substrates is needed to reveal their mechanism of action, but high-throughput methods to determine glycan specificity of neuraminidases are limited. Here we use two glycan labeling reactions to monitor neuraminidase activity toward glycan substrates. While both periodate oxidation and aniline-catalyzed oxime ligation (PAL) and galactose oxidase and aniline-catalyzed oxime ligation (GAL) can be used to monitor neuraminidase activity toward glycans in microtiter plates, only GAL accurately measured neuraminidase activity toward glycans displayed on a commercial glass slide microarray. Using GAL, we confirm known linkage specificities of three pneumococcal neuraminidases and obtain new information about underlying glycan specificity.
authors.affiliation: Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Janet E
authors.initials: JE
authors.lastname: McCombs
authors.affiliation: Division of Cardiology, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Jason P
authors.initials: JP
authors.lastname: Diaz
authors.affiliation: Division of Cardiology, Department of Internal Medicine, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA.
authors.forename: Kevin J
authors.initials: KJ
authors.lastname: Luebke
authors.affiliation: Department of Biochemistry, University of Texas Southwestern Medical Center, Dallas, TX 75390, USA. Electronic address: jennifer.kohler@utsouthwestern.edu.
authors.forename: Jennifer J
authors.initials: JJ
authors.lastname: Kohler
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D046228:Microarray Analysis / Q000379:methods*; D009439:Neuraminidase / Q000235:genetics / Q000378:metabolism*; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism*; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013194:Staining and Labeling; D013296:Streptococcus pneumoniae / Q000201:enzymology* / Q000235:genetics; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D011134:Polysaccharides; D011994:Recombinant Proteins; D009439:Neuraminidase
keywords: Chemoenzymatic labeling; Microarray; Neuraminidase; Sialic acids