title: Glycosylation of Rapana thomasiana hemocyanin. Comparison with other prosobranch (gastropod) hemocyanins.
abstract: The carbohydrate content and composition of hemocyanins (Hcs) of three prosobranchs (gastropods), Rapana thomasiana, Megathura crenulata and Haliotis tuberculata, were compared. The analyses were performed by gas-liquid chromatography after methanolysis, re-N-acetylation and trimethylsilylation. The two structural subunits of R. thomasiana Hc, RtH1 and RtH2, both showed 2.6% (w/w) carbohydrate content with very similar monosaccharide composition, indicative for N-glycosylation. The two isoforms of M. crenulata Hc (KLH), KLH1 and KLH2, on the other hand, definitely differed in glycosylation: KLH2 (3.4% carbohydrate, w/w) comprised relatively less mannose and more N-acetylgalactosamine than KLH1 (3.0% carbohydrate, w/w), in agreement with the fact that O-glycosylation has been observed in a functional unit (FU) of KLH2. For the Hc of the abalone H. tuberculata, with 4.5% (w/w) carbohydrate, appreciable amounts of 3-O-methyl-d-mannose and 3-O-methyl-d-galactose were detected, showing that the occurrence of methylated sugars is not restricted to the Hcs of pulmonates. From the structural subunit RtH2 of Rapana Hc the FUs RtH2-b and RtH2-d were isolated. On the basis of amino acid sequence analysis and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) of the respective native and PNGase-F-treated glycopeptides, one N-glycosylation site was found for each FU. This site was located at Asn-405 for RtH2-b and at Asn-394 for RtH2-d; the carbohydrate moiety corresponded to GlcNAc2Man6 and GlcNAc2Man5, respectively. A comparison was made with the N-glycosylation sites of other FUs of Rapana Hc.
authors.affiliation: Institute of Organic Chemistry, Bulgarian Academy of Sciences, Akad. G. Bonchev-Str. bl. 9, Sofia 1113, Bulgaria. idakieva@orgchm.bas.bg
authors.forename: Krassimira
authors.initials: K
authors.lastname: Idakieva
authors.forename: Stanka
authors.initials: S
authors.lastname: Stoeva
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Voelter
authors.forename: Constant
authors.initials: C
authors.lastname: Gielens
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D002240:Carbohydrate Sequence; D002849:Chromatography, Gas; D006020:Glycopeptides / Q000737:chemistry / Q000378:metabolism; D006031:Glycosylation; D006433:Hemocyanins / Q000378:metabolism*; D008969:Molecular Sequence Data; D012908:Snails / Q000737:chemistry* / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D006020:Glycopeptides; D006433:Hemocyanins