Amino acid sequence and glycosylation of functional unit RtH2-e from Rapana thomasiana (gastropod) hemocyanin
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Updated: Oct 2, 2026

Targeting Cysteine Thiols for in Vitro Site-specific Glycosylation of Recombinant Proteins
Published on: October 4, 2017
Stanka Stoeva1, Krasimira Idakieva, Christian Betzel
1Department of Physical Biochemistry, Institute of Physiological Chemistry, University of Tuebingen, Hoppe-Seyler-Strasse 4, D-72076 Tuebingen, Germany. stanka.stoeva@uni-tuebingen.de
Abstract:
The complete amino acid sequence of Rapana thomasiana hemocyanin functional unit RtH2-e was determined by direct sequencing and matrix-assisted laser desorption ionization mass spectrometry of peptides obtained by cleavage with EndoLysC proteinase, chymotrypsin, and trypsin. The single-polypeptide chain of RtH2-e consists of 413 amino acid residues and contains two consensus sequences NXS/T (positions 11-19 and 127-129), potential sites for N-glycosylation. Monosaccharide analysis of RtH2-e revealed a carbohydrate content of about 1.1% and the presence of xylose, fucose, mannose, and N-acetylglucosamine, demonstrating that only N-linked carbohydrate chains of high-mannose type seem to be present. On basis of the monosaccharide composition and MALDI-MS analysis of native and PNGase-F-treated chymotryptic glycopeptide fragment of RtH2-e the oligosaccharide Man(5)GlcNAc(2), attached to Asn(127), is suggested. Multiple sequence alignments with other molluscan hemocyanin e functional units revealed an identity of 63% to the cephalopod Octopus dofleini and of 69% to the gastropod Haliotis tuberculata. The present results are discussed in view of the recently determined X-ray structure of the functional unit g of the O. dofleini hemocyanin.
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