Characterization of the N-glycans of female Angiostrongylus cantonensis worms.
Glycoconjugates play a crucial role in the host-parasite relationships of helminthic infections, including angiostrongyliasis. It has previously been shown that the antigenicity of proteins from female Angiostrongylus cantonensis worms may depend on their associated glycan moieties. Here, an N-glycan profile of A. cantonensis is reported. A total soluble extract (TE) was prepared from female A. cantonensis worms and was tested by western blot before and after glycan oxidation or N- and O-glycosidase treatment. The importance of N-glycans for the immunogenicity of A. cantonensis was demonstrated when deglycosylation of the TE with PNGase F completely abrogated IgG recognition. The TE was also fractionated using various lectin columns [Ulex europaeus (UEA), concanavalin A (Con A), Arachis hypogaea (PNA), Triticum vulgaris (WGA) and Lycopersicon esculentum (LEA)], and then each fraction was digested with PNGase F. Released N-glycans were analyzed with matrix-assisted laser desorption ionization (MALDI)-time-of-flight (TOF)-mass spectrometry (MS) and MALDI-TOF/TOF-MS/MS. Complex-type, high mannose, and truncated glycan structures were identified in all five fractions. Sequential MALDI-TOF-TOF analysis of the major MS peaks identified complex-type structures, with a α1-6 fucosylated core and truncated antennas. Glycoproteins in the TE were labeled with BodipyAF558-SE dye for a lectin microarray analysis. Fluorescent images were analyzed with ProScanArray imaging software followed by statistical analysis. A total of 29 lectins showed positive binding to the TE. Of these, Bandeiraea simplicifolia (BS-I), PNA, and Wisteria floribunda (WFA), which recognize galactose (Gal) and N-acetylgalactosamine (GalNAc), exhibited high affinity binding. Taken together, our findings demonstrate that female A. cantonensis worms have characteristic helminth N-glycans.
Laboratório de Parasitologia Molecular, Instituto de Pesquisas Biomédicas and Laboratório de Biologia Parasitária, Faculdade de Biociências da Pontifícia Universidade Católica do Rio Grande do Sul (PUCRS), Porto Alegre, RS 90060-900, Brazil; Institute for Glycomics, Griffith University, Gold Coast Campus, Queensland, 4222, Australia. Electronic address: caroldmv@yahoo.com.br.
Carolina M
CM
Veríssimo
Laboratório de Parasitologia Molecular, Instituto de Pesquisas Biomédicas and Laboratório de Biologia Parasitária, Faculdade de Biociências da Pontifícia Universidade Católica do Rio Grande do Sul (PUCRS), Porto Alegre, RS 90060-900, Brazil.
Alessandra L
AL
Morassutti
Institute for Glycomics, Griffith University, Gold Coast Campus, Queensland, 4222, Australia.
Mark
M
von Itzstein
Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London, SW7 2AZ, United Kingdom.
Grigorij
G
Sutov
Institute for Glycomics, Griffith University, Gold Coast Campus, Queensland, 4222, Australia.
Lauren
L
Hartley-Tassell
Institute for Glycomics, Griffith University, Gold Coast Campus, Queensland, 4222, Australia.
Sarah
S
McAtamney
Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London, SW7 2AZ, United Kingdom.
Anne
A
Dell
Department of Life Sciences, Faculty of Natural Sciences, Imperial College London, London, SW7 2AZ, United Kingdom.
Stuart M
SM
Haslam
Laboratório de Parasitologia Molecular, Instituto de Pesquisas Biomédicas and Laboratório de Biologia Parasitária, Faculdade de Biociências da Pontifícia Universidade Católica do Rio Grande do Sul (PUCRS), Porto Alegre, RS 90060-900, Brazil.
Carlos
C
Graeff-Teixeira
D017185:Angiostrongylus cantonensis / Q000737:chemistry* / Q000276:immunology; D000818:Animals; D005260:Female; D006026:Glycoside Hydrolases / Q000378:metabolism; D037102:Lectins / Q000378:metabolism; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000378:metabolism; D010504:Periodic Acid / Q000378:metabolism; D011134:Polysaccharides / Q000276:immunology* / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
D037102:Lectins; D011134:Polysaccharides; D010504:Periodic Acid; C009288:metaperiodate; D006026:Glycoside Hydrolases; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase
Angiostrongylus cantonensis; Glycans; Lectin array; Mass spectrometry