Glycosylation sites, determination

The equivalent of the tryptic fragment of human transferrin receptor has been expressed in Chinese hamster ovary cells and its structure determined at a resolution of 0.32 nm (Lawrence et ah, 1999). The asymmetric unit of the crystals contains four transferrin receptor dimers. Interpretable electron density is found for the entire tryptic fragment except for Arg-121 at the amino terminus, and density is also seen for the first N-acetylglucosamine residue at each of the N-glycosylation sites. The transferrin receptor monomer is made up of three distinct domains, organized such that the dimer is butterfly shaped (Figure 5.10, Plate 7). The likely orientation of the dimer with respect to the plasma membrane has been assigned on the basis of the... 

Cuyckens F and Claeys M. 2005. Determination of the glycosylation site in llavonoid mono-O-glycosides by collision-induced dissociation of electrospray-generated deprotonated and sodiated molecules. J Mass Spectrom 40(3) 364-372. 

Fig. 2. Primary structure of hamster PrP (Stahl et al., 1993). The first 22 residues at the N-terminus are the signal sequence. PrPc is completely digested by proteinase K, whereas the N-terminal sequence of PrPSc to residue 89 (arrow, closed head) is digested. — CHO indicates the glycosylation sites at residues 181 and 197 Gpi the glycosylpho-sphatidylinositol anchor at 231 and the N-terminal octarepeats. In one case of human prion disease, a stop codon was found at 145 (arrow, open head) (Kitamoto et al., 1993). HI, H2, H3, and H4 denote the predicted a-helices (Huang et al, 1994, 1996), and A-C denote the a-helices and SI, S2 the /(-strands determined by solution NMR (James et al, 1997).

Davis, B.D. and Brodbelt, J.S., Determination of the glycosylation site of flavonoid monogluco-sides by metal complexation and tandem mass spectrometry, J. Am. Chem. Soc. Mass Spectrom.,... 

G. J. Rademaker, J. Haverkamp, and J. E. Thomas-Oates, Determination of glycosylation sites in O-linked glycopeptides A sensitive mass spectrometric protocol, Org. Mass Spectrom., 28 (1993) 1536-1541. 

E. Mirgorodskaya, H. Hassan, H. Clausen, and P. Roepstorff, Mass spectrometric determination of O-glycosylation sites using P-elimination and partial acid hydrolysis, Anal. Chem., 73 (2001) 1263-1269. 

B. Macek, J. Hofsteenge, and J. Peter-Katalinic, Direct determination of glycosylation sites in O-fucosylated glycopeptides using nano-electrospray quadrupole time-of-flight mass spectrometry, Rapid Commun. Mass Spectrom., 15 (2001) 771-777. 

M. Svoboda, M. Przyblski, J. Schreurs, A. Miyajima, K. Hogeland, and M. Deinzer, Mass spectrometric determination of glycosylation sites and oligosaccharide composition of insect-expressed mouse interleukin-3, J. Chromatogr., 562 403-419 (1991). 

Stellacyanin is also an important example of a heavily glycosylated protein for which the structure has been determined without its glyco components. It demonstrated that the carbohydrate moieties have virtually no effect on the folding topology of the polypeptide core of this particular glycoprotein. One of the three glycosylation sites in... 

J. Gonzalez, T. Takao, H. Hori, V. Besada, R. Rodrignez, G. Padron, and Y. Shimonishi, A method for determination of N-glycosylation sites in glycoproteins by collision-indnced dissociation analysis in fast atom bombardment mass spectrometry Identification of the positions of carbohydrate-linked asparagine in recombinant a-amylase by treatment with peptide-A-glycosidase F in 0-labeled water, A flZ. Biochem. 205 (1992), 151-158. 

NMR studies on purine nucleosides have been undertaken to assign the site of glycosylation, to determine the anomeric Configuration, to elucidate the sugar puckering and to determine the synjanti ratio of the location of the bases. Rare tautomeric forms and protonation sites were also studied. 

The presence of the glycan (light blue) on ribonuclease B reduces the amide proton/deuteiium exchange rates compared with ribonuclease A for extensive regions of the peptide backbone (shown in red) both local to and remote from the glycosylation site [47]. The glycoprotein structure is based on the crystal structure of ribonuclease B [53] and one of the structures of MangGlcNAc2 determined by NMR and molecular dynamics. (Reprinted from [44] with permission from Elsevier), [54]... 

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