Glycoconjugate Journal

, Volume 27, Issue 1, pp 159–169 | Cite as

N-glycan moieties of the crustacean egg yolk protein and their glycosylation sites

  • Ziv Roth
  • Shmuel Parnes
  • Simy Wiel
  • Amir Sagi
  • Nili Zmora
  • J. Sook Chung
  • Isam Khalaila
Article

Abstract

Vitellogenin (Vg) is the precursor of the egg yolk glycoprotein of crustaceans. In the prawn Macrobrachium rosenbergii, Vg is synthesized in the hepatopancreas, secreted to the hemolymph, and taken up by means of receptor-mediated endocytosis into the oocytes. The importance of glycosylation of Vg lies in its putative role in the folding, processing and transport of this protein to the egg yolk and in the fact that the N-glycan moieties could provide a source of carbohydrate during embryogenesis. The present study describes, for the first time, the structure of the glycan moieties and their sites of attachment to the Vg of M. rosenbergii. Bioinformatics analysis revealed seven putative N-glycosylation sites in M. rosenbergii Vg; two of these glycosylation sites are conserved throughout the Vgs of decapod crustaceans from the Pleocyemata suborder (N 159 and N 660). The glycosylation of six putative sites of M. rosenbergii Vg (N 151, N 159, N , 168 N , 614 N 660 and N 2300) was confirmed; three of the confirmed glycosylation sites are localized around the N-terminally conserved N-glycosylation site N 159. From a theoretical three-dimensional structure, these three N-glycosylated sites N 151, N 159, and N 168 were localized on the surface of the Vg consensus sequence. In addition, an uncommon high mannose N-linked oligosaccharide structure with a glucose cap (Glc1Man9GlcNAc2) was characterized in the secreted Vg. These findings thus make a significant contribution to the structural elucidating of the crustacean Vg glycan moieties, which may shed light on their role in protein folding and transport and in recognition between Vg and its target organ, the oocyte.

Keywords

Crustacea Glucose cap Glycosylation sites N-glycan Vitellogenin 

Abbreviations

ACN

acetonitrile

DIG

digoxigenin

DSA

Datura stramonium agglutinin

DTT

dithiothreitol

ER

endoplasmic reticulum

Glc

glucose

GlcNAc

N-acetylglucosamine

GII

glucosidase II

GNA

Galanthus nevallis agglutinin

GU

glucose unit

HDL

high density lipoprotein

Hex

hexose

HexNAc

N-acetylhexosamine

HPLC

high performance liquid chromatography

JBM

jack bean α-mannosidase

MAA

Maackia amurensis agglutinin

Man

mannose

MALDI

matrix-assisted laser desorption/ionization

PNA

peanut (Arachis hypogaea) agglutinin

PNGase F

peptide-N-glycosidase F

SDS-PAGE

sodium dodecyl sulfate polyacrylamide gel electrophoresis

SNA

Sambucus nigra agglutinin

TFA

trifluoroacetic acid

Vg

vitellogenin

Notes

Acknowledgments

We thank Ayana Benet-Perlberg and the team from the Dor experimental station of the Ministry of Agriculture for providing animal culture facilities, Mr. Tomer Ventura for his technical assistance and Ms. Lilah Glazer for her help in generating the 3D model. This work was supported by an internal grant from Ben Gurion University of the Negev; I. K. is the incumbent of MAOF fellowship from the Israel Council for Higher Education, Planning and Budgeting Committee.

Supplementary material

10719_2009_9268_MOESM1_ESM.pdf (193 kb)
ESM 1 (PDF 193 kb)

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Copyright information

© Springer Science+Business Media, LLC 2009

Authors and Affiliations

  • Ziv Roth
    • 1
    • 2
    • 3
  • Shmuel Parnes
    • 2
    • 3
  • Simy Wiel
    • 2
    • 3
  • Amir Sagi
    • 2
    • 3
  • Nili Zmora
    • 4
  • J. Sook Chung
    • 4
  • Isam Khalaila
    • 1
  1. 1.Department of Biotechnology EngineeringBen-Gurion University of the NegevBeer-ShevaIsrael
  2. 2.Department of Life SciencesBen-Gurion University of the NegevBeer-ShevaIsrael
  3. 3.The National Institute for Biotechnology in the NegevBen-Gurion University of the NegevBeer-ShevaIsrael
  4. 4.Center of Marine Biotechnology, University of Maryland Biotechnology InstituteBaltimoreUSA

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