A facile synthesis of Nγ-glycosyl asparagine conjugates and short N-linked glycopeptides

Article abstract of DOI:10.1080/07328303.2011.633723

Both free and protected glycosyl azides were efficiently coupled to the side chain of aspartate by the Staudinger reaction for the synthesis of N ^γ-glycosyl asparagine conjugates and short N-linked glycopeptides that can be employed to construct complex N-linked glycopeptides. In the process, a facile two-step protocol was developed for free glycosyl azide synthesis, which includes reported direct transformation of free oligosaccharides into glycosyl amines through reaction with ammonium bicarbonate and then stereospecific diazo transfer reaction to convert glycosyl amines to glycosyl azides.

Full text of DOI:10.1080/07328303.2011.633723

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A Facile Synthesis of N γ-Glycosyl
Asparagine Conjugates and Short N-
Linked Glycopeptides
Jie Xue a , Min Guo a , Guofeng Gu b & Zhongwu Guo a b
a Department of Chemistry , Wayne State University , Detroit , MI ,
48202 , USA
b National Glycoengineering Research Center , Shandong University ,
Ji’nan , 250100 , China
Published online: 07 Feb 2012.
To cite this article: Jie Xue , Min Guo , Guofeng Gu & Zhongwu Guo (2012) A Facile Synthesis of
N γ-Glycosyl Asparagine Conjugates and Short N-Linked Glycopeptides, Journal of Carbohydrate
Chemistry, 31:2, 105-113, DOI: 10.1080/07328303.2011.633723
To link to this article: http://dx.doi.org/10.1080/07328303.2011.633723
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Journal of Carbohydrate Chemistry, 31:105–113, 2012
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Copyright Taylor & Francis Group, LLC
ISSN: 0732-8303 print / 1532-2327 online
DOI: 10.1080/07328303.2011.633723
A Facile Synthesis of
Nγ-Glycosyl Asparagine
Conjugates and Short
N-Linked Glycopeptides
Jie Xue,1 Min Guo,1 Guofeng Gu,2 and Zhongwu Guo3
1Department of Chemistry, Wayne State University, Detroit, MI 48202, USA
2National Glycoengineering Research Center, Shandong University,
Ji’nan 250100, China
3Department of Chemistry, Wayne State University, Detroit, MI 48202, USA;
and National Glycoengineering Research Center, Shandong University,
Ji’nan 250100, China
Both free and protected glycosyl azides were efficiently coupled to the side chain of
γ
aspartate by the Staudinger reaction for the synthesis of N -glycosyl asparagine con-
jugates and short N-linked glycopeptides that can be employed to construct complex
N-linked glycopeptides. In the process, a facile two-step protocol was developed for free
glycosyl azide synthesis, which includes reported direct transformation of free oligosac-
charides into glycosyl amines through reaction with ammonium bicarbonate and then
stereospecific diazo transfer reaction to convert glycosyl amines to glycosyl azides.
Keywords Glycosyl asparagine; Glycopeptide; Glycosyl azide; Thioester; Staudinger
reaction
INTRODUCTION
N-Glycosylation of proteins, that is, attachment of glycans to the asparagine
side chain of peptide backbones through the glycan reducing end, is one of
the most common posttranslational modifications.[1] N-Glycoproteins and gly-
copeptides play an important role in various biological processes.[2] However,
Received August 29, 2011; accepted October 17, 2011.
Address correspondence to Zhongwu Guo, Department of Chemistry, Wayne State Uni-
versity, 5101 Cass Avenue, Detroit, MI 48202, USA; and National Glycoengineering
Research Center, Shandong University, 29-1 Shanda Nan Lu, Ji’nan 250100, China.
E-mail: zwguo@chem.wayne.edu
105

J. Xue et al.
106
it is very difficult, if not impossible, to isolate homogenous glycoproteins and
glycopeptides from natural sources because of the inherent microheterogene-
ity. Therefore, access to structurally homogenous and defined glycoforms of
glycoproteins and glycopeptides needs to rely on chemical/chemoenzymatic
synthesis. For this purpose, many methods have been developed in the past
decades,[3,4] despite various significant challenges.[5]
A common problem in glycopeptide synthesis is the incompatibility be-
tween peptide and carbohydrate chemistries. Currently, there are two general
strategies to deal with this problem besides enzymatic synthesis.[6] One de-
pends on the coupling of full-length synthetic carbohydrates and peptides,[7,8]
which is a highly convergent strategy but is often hampered by the poor yield
and selectivity of reactions employed to form the amino acid–glycan linkage.
A more widely adopted strategy is based on the use of glycosyl amino acids,
derived from coupling of oligosaccharides and amino acids, as key building
blocks for constructing the desired glycopeptides by stepwise solid-phase and
solution-phase elongation of the peptide chain.[9,10] Some potential issues with
the latter strategy are associated with the final stage of carbohydrate depro-
tection, as well as the harsh conditions utilized to detach glycopeptides from
resins in solid-phase glycopeptide synthesis. To address these potential issues,
our group has developed a new synthetic strategy called “solution-phase gly-
copeptide synthesis with solid-state workup,”[11–14] which uses free glycosyl
asparagine conjugates as key building blocks. For either the traditional solid-
and solution-phase synthesis strategy or the solution-phase synthesis with
solid-state workup strategy, a critical step is to access glycosyl asparagine
conjugates, which are usually synthesized by coupling glycosyl amines to an
aspartate side chain carboxyl group.[10] A major problem associated with
these syntheses is glycan-reducing end-anomeric epimerization of the glyco-
syl amines involved.
This work aimed to develop a stereoselective approach for the prepara-
tion of glycosyl asparagine conjugates applicable to the synthesis of N-linked
glycopeptides either by the solution-phase synthesis with solid-state workup
strategy or by the conventional solution- and solid-phase glycopeptide syn-
thesis with glycosyl amino acid building blocks. In this context, we became
interested in chemoselective ligation methods using the Staudinger reaction,
which has been proved to be useful for the creation of amide bonds in various
applications.[15–17] For example, Raines and co-workers[18,19] developed a new
amino acid coupling method based on stereospecific reactions of phosphinoth-
ioesters 1 and azides as outlined in Scheme 1. Kiessling[20] and Bernardi[21]
used the same or similar reaction to couple glycosyl azides and amino acids.
Several other amino acid–glycan ligation methods[22–24] also involved a simi-
lar mechanism. An important property of this coupling reaction is that it is
stereospecific, and thus glycan-reducing end-anomeric epimerization can be
avoided, which will simplify the purification of resulting glycosyl amino acids.

Synthesis of N-Glycopeptides
107
Scheme 1: Peptide synthesis via the ligation reaction between peptide phosphinothioester
and azide.
Moreover, glycosyl azides are commonly used intermediately in the synthesis
of N-linked glycopeptides.[25]
RESULTS AND DISCUSSION
Many free oligosaccharides are commercially available. To employ them in the
synthesis of asparagine conjugates and glycopeptides, we developed a con-
venient and efficient method to directly transform free oligosaccharides to
glycosyl azides. As shown in Scheme 2, first, we can convert a free oligosaccha-
ride, such as maltotriose 5, to the corresponding glycosyl amine (6),[26] which
is then converted to glycosyl azide (7) via stereospecific diazo transfer reac-
tion.[27,28] The reaction between 5 and NH4HCO3 went smoothly to give more
polar 6 as revealed by TLC. After the reaction mixture was condensed to re-
move NH4HCO3 and solvent completely, the crude 6 was dissolved in DMF and
reacted with freshly prepared N3Tf with CuSO4 as catalyst, which resulted in
glycosyl azide 7 in a 50% isolated yield upon precipitation with diethyl ether.
As the reaction was clean but the reaction yield was relatively low, we assumed
that it was because of the poor solubility of the glycosyl amine intermediate
in DMF. To address this problem, we switched the solvent to DMSO, which
1
improved the reaction yield to 83%. H NMR spectrum of 7 clearly showed
the β-configuration of its glycosyl azide linkage. Evidently, β-linked glycosyl
amine 6 was formed almost exclusively in the previous step, as reported in the
literature.[26]
Scheme 2: Direct conversion of free oligosaccharide to glycosyl azide.

J. Xue et al.
108
Thereafter, the γ-diphenylphosphinothioester 8 of aspartic acid was pre-
pared according to the reported procedure,[18,19] and its reaction with 7 was
probed in THF/H2O (3:1). However, this reaction only resulted in a small
amount of the coupling product 9, while the majority of 7 was reduced to form
glycosyl amine 6 and free glycan 5, which might result from the hydrolysis
of involved reactive intermediate like 2. To eliminate this side reaction, we
switched the solvent to dry DMF. Under such condition, the coupling reaction
between 7 and 8 proceeded smoothly to afford the desired glycosyl asparagine
conjugate 9 in a 55% yield (Sch. 3) after precipitation and column chromato-
graph. It was further observed that the coupling reaction was stereospecific to
provide the β-linked conjugate 9 only. The spectroscopic data of 9 were identi-
cal to that of the authentic sample prepared by a different method.[13]
Scheme 3: Coupling reaction between free glycosyl azide and amino acid
phosphinothioester.
Next, we studied the reaction between 8 and 10,[12] which is a partial
structure of N-linked glycan core with a fucosyl branch. The synthesis of this
oligosaccharide and the core of N-linked glycans, as well as their asparagine
conjugates, has already been reported in the literature.[29,30] It was proved that
the sterically demanding glycan 10 could be efficiently coupled with 8 to afford
the conjugate product 11 in a 70% yield (Sch. 4). Again, we found that the re-
action was stereospecific to produce only the β-anomer. The NMR spectra of 11
were identical to that reported previously.[12]
Scheme 4: Coupling reaction between protected glycosyl azide and amino acid
phosphinothioester.
Encouraged by the above results, we finally explored the ligation of glycans
with peptides by the same strategy. The reaction between 10 and dipeptide 12
proceeded smoothly to afford the coupling product 13 (70%, Sch. 5), which is

Synthesis of N-Glycopeptides
109
Scheme 5: Coupling reactions between glycosyl azides and peptide phosphinothioesters.
useful for convergent glycopeptide synthesis, namely, using simple glycopep-
tides as building blocks for the assembly of complex glycopeptides. However,
the reaction of 10 with tripeptide 14 provided only the corresponding glycosyl
amine rather than the coupling product, whereas the reaction of 14 with 15
gave the coupling product 16 in a good yield (72%). These results may sug-
gest that if both the glycan and peptide moieties are sterically hindered, the
increased steric hindrance may obstruct the intramolecular acyl transfer re-
action of intermediates like 2; thus, only the reduction and hydrolysis product
was formed.
In summary, we have developed a simple, efficient, and potentially broadly
applicable method for the preparation of glycosyl azides from free oligosac-
charides via two well-established, stereoselective reaction steps. Glycosyl
azides carrying both free and protected oligosaccharides reacted stereospecif-
ically with γ-diphenylphosphinothioester of aspartic acid to provide glyco-
syl asparagine conjugates that are useful for glycopeptide assembly by the
solution-phase synthesis with solid-state workup strategy or by the conven-
tional solution- and solid-phase synthesis strategy, respectively. On the other
hand, we observed that the reaction of 8 with peracetylated 7 gave only a very
low yield (∼10%) of the desired glycosyl asparagine conjugate, while the ma-
jor product was the corresponding glycosyl amine. It suggested that the in-
tramolecular acyl transfer reaction of intermediate 2 might be suppressed by
the electron-deficient and anion-stabilizing acylated glycan; as a result, hy-
drolysis of 2 prevailed. We have also shown that glycosyl azides can be cou-
pled with short peptides to form glycopeptides that may be useful as building
blocks for the construction of more complex N-glycopeptides. However, it seems
that the intramolecular acyl transfer reaction is rather sensitive to the sub-
strate structure, because while dipeptide 12 was successfully coupled to 10, a
slightly more sterically demanding tripeptide 14 was not, though 14 could be
successfully coupled with less sterically hindered glycosyl azide 15. Our con-
clusion is that electron-rich and less sterically hindered glycans and peptides

J. Xue et al.
110
are more likely to undergo intramolecular acyl transfer reaction via interme-
diate 2 to form the desirable glycan–amino acid coupling products. Overall, we
have introduced herein an alternative method for the preparation of glycosyl
asparagine conjugates and short glycopeptides.
EXPERIMENTAL
General Methods
NMR spectra were recorded with a Varian-400 FT NMR spectrometer. Cou-
pling constants (J) are reported in Hertz (Hz). Optical rotations were recorded
with a Perkin-Elmer 241 polarimeter. High-resolution mass spectra (HRMS)
and MS were recorded with a Kratos MS-25RFA mass spectrometer using fast
atom bombardment (FAB) conditions. Thin layer chromatography (TLC) was
performed on silica gel GF254 with detection by charring with 2% H2SO4/EtOH.
Anhydrous solvents and other reagents were purchased from Aldrich and used
without further purification. Glycosyl azide 10 was prepared according to a
method previously reported by this lab.[12]
α-D-Glucopyranosyl-(1→4)-α-D-glucopyranosyl-(1→4)-
α-D-glucopyranosyl Azide (7)
After 5 (101 mg, 0.2 mmol) and NH4HCO3 (0.2 mol) dissolved in saturated
NH4OH aq. solution (5 mL) was stirred at 42◦C for 2 d and TLC showed com-
plete reaction, the mixture was concentrated under vacuum to afford glycosyl
amine 6 as a white solid. The crude product was dissolved in 2 mL of DMSO,
and to the solution was added CuSO4 (cat. amount) and freshly prepared N3Tf
(2 eq.) in CH2Cl2. The reaction mixture was stirred at rt for 1 d and then
concentrated. To the remaining DMSO solution was added excessive Et2O
(20 mL). The resultant glycosyl azide 7 was collected as a white precipitate,
1
which was practically pure as shown by H NMR and was then directly used
in the next step of reaction. The NMR and MS spectra of 7 were identical to
that of the reported.[13]
Glycosyl Azide Ligation with Aspartate or Peptide
(General Procedure)
A solution of aspartic acid thioester 8, dipeptide thioester 12, or tripep-
tide thioester 14 (0.02 mmol) and glycosyl azide 7 or 10 (0.02 mmol) dissolved
in dry DMF (2.0 mL) was stirred at rt overnight under an argon atmosphere.
TLC showed that the reaction was completed. Then, Et2O (20 mL) was added
to the reaction mixture, and the resultant precipitates were collected and sub-
jected to purification by flash column chromatography or preparative TLC to

Synthesis of N-Glycopeptides
111
afford glycosyl asparagine conjugates and glycopeptides 9, 11, 13, and 16, re-
spectively. The spectroscopic data of 9 and 11 were identical to that reported
previously,[12,13] while the spectroscopic and other physical data of new com-
pounds involved in the syntheses are listed below.
tert-Butyl γ-diphenylphosphinothio-Nα-fluorenylmethyloxycarbonyl-
L-aspartate 8
[α] = +30.5 (1.0, CHCl3); 1H NMR (CDCl3, 400 MHz): 7.74 (d, J = 7.6 Hz,
2H), 7.57 (d, J = 7.2 Hz, 2H), 7.40–7.23 (m, 14H), 5.60 (d, J = 8.4 Hz, 1H,
NH), 4.44 (dt, J = 8.0, 4.4 Hz, 1H, α-H), 4.34–4.32 (m, 2H, Fmoc), 4.20 (t, J =
7.2 Hz, 1H, Fmoc), 3.51–4.49 (m, 2H, PCH2S), 3.14 (dd, J = 16.4, 4.4 Hz, 1H,
β-H of Asp), 3.04 (dd, J = 16.4, 4.4 Hz, 1H, β-H of Asp), 1.42 (s, 9H, t-Bu); 13
C
NMR (CDCl3, 100 MHz): 169.3, 155.9, 143.9, 141.4, 133.0, 132.9, 132.8, 129.4,
128.8, 128.7, 127.9, 127.2, 125.3, 120.1, 83.0, 67.3, 51.4, 47.3, 45.4, 28.0, 26.0,
25.8; 31P NMR (CDCl3): −15.06; FAB HRMS: calcd. for C36H37NO5PS (M +
H+): 626.2130; found: 626.2122.
tert-Butyl Nα-fluorenylmethyloxycarbonyl-glycinyl-γ-diphenylphosphinothio-
L-aspartate 12
1H NMR (CDCl3, 400 MHz): 7.72 (d, J = 7.2 Hz, 2H), 7.57 (d, J = 6.8 Hz,
2H), 7.37–7.23 (m, 14H), 6.69 (d, J = 7.6 Hz, 1H, NH), 5.40 (br, 1H, NH), 4.64
(dt, J = 7.6, 4.4 Hz, 1H, α-H), 4.40–4.26 (m, 2H, Fmoc), 4.21 (t, J = 7.2 Hz,
1H, Fmoc), 3.87–3.84 (m, 2H, Gly-CH2), 3.46–3.44 (m, 2H, PCH2S), 3.12 (dd,
J = 16.4, 4.4 Hz, 1H, β-H of Asp), 3.04 (dd, J = 16.4, 4.4 Hz, 1H, β-H of Asp),
1.41 (s, 9H, t-Bu); 31P NMR (CDCl3): −11.87. FAB MS: calcd. for C36H36NO5PS
(M + H+): 625; found 625.
tert-Butyl N-fluorenylmethyloxycarbonyl-glycinyl-Nγ-{2-acetamido-2-deoxy-
β-D-glucopyranosyl-(1→4)-[2,3,4-O-tribenzyl-α-L-fucopyranosyl-(1→6)]-
2-acetamido-3-O-benzyl-2-deoxy-β-D-glucopyranosyl}-L-asparaginate 13
[α] = −18.0 (0.5, CHCl3); 1H NMR (CD3OD, 400 MHz): 7.75 (d, J = 7.6 Hz,
2H, Fmoc), 7.62 (d, J = 6.0 Hz, 2H, Fmoc), 7.36–7.21 (m, 24H), 5.01 (d, J =
10.8 Hz, 1H, H-1), 4.91 (d, J = 9.6 Hz, 1H, H-1), 4.65 (d, J = 8.4 Hz, 1H, H-1),
4.18 (t, J = 6.8 Hz, 1H, Fmoc), 2.71 (dd, J = 16.4, 5.2 Hz, 1H, β-H of Asp), 2.61
(dd, J = 16.4, 5.2 Hz, 1H, β-H of Asp), 1.95 (s, 3H, Ac), 1.87 (s, 3H, Ac), 1.39 (s,
9H, t-Bu), 1.08 (d, J = 6.4 Hz, 3H, Fuc-Me); FAB MS: calcd. for C75H90N5O20
(M + H+): 1381; found 1381.
tert-Butyl N-fluorenylmethyloxycarbonyl-glycinyl-Nγ-{2-acetamino-4,6-
O-benzylidene-2-deoxy-β-D-glucopyranosyloxyethyl}-L-
asparaginyl-glycinate 16
1
[α] = −5.2 (1.0, CHCl3); H NMR (CDCl3, 400 MHz): 7.82 (q, 2H, Phth),
7.72 (d, J = 7.6 Hz, 2H, Fmoc), 7.69 (q, 2H, Phth), 7.57–7.25 (m, 11H), 6.15 (br,

J. Xue et al.
112
1H, NH), 5.59 (br, 1H, NH), 5.51 (s, 1H, benzylidene), 5.25 (d, J = 8.4 Hz, 1H,
H-1), 4.67 (dt, J = 4.0, 6.8 Hz, 1H, α-H of Asp), 4.59 (dd, J = 10.0, 8.8 Hz, 1H,
H-3), 4.41–4.31 (m, 3H, Fmoc-CH2 and H-5), 4.21–4.19 (m, 2H, Fmoc-CH and
H-2), 3.91 (d, J = 5.6 Hz, 2H, α-H of Gly), 3.87 (d, J = 5.6 Hz, 2H, α-H of Gly),
3.79–3.71 (m, 2H, H-6 and linker-H), 3.64 (s, 3H, Me), 3.62–3.58 (m, 3H, H-4,
H-6, linker-H), 3.29–3.26 (m, 2H, linker-H), 2.73 (br, 1H, OH), 2.59 (dd, J =
15.2, 3.2 Hz, 1H, β-H of Asp), 2.36 (dd, J = 15.2, 2.8 Hz, 1H, β-H of Asp); FAB
HRMS (m/e): calcd. for C47H47N5NaO14 (M + Na+): 928.3017; found 928.3052.
ACKNOWLEDGEMENT
We thank Mizutani Foundation for Glycoscience, Japan, and NSF, USA (CHE-
10538480), for the research awards to support this work.
REFERENCES
1. Cummings, R.D. Synthesis of asparagine-linked oligosaccharides: pathways, ge-
netics, and metabolic regulation. In Glycoconjugates, H.J. Allen and E.C. Kisailus, Eds.
Marcel Dekker: New York, 1992, 333–358.
2. Helenius, A.; Aebi, M. Intracellular functions of N-linked glycans. Science 2001,
291, 2364–2369.
3. Seitz, O. Glycopeptide synthesis and the effects of glycosylation on protein struc-
ture and activity. Chem. Bio. Chem. 2000, 1, 214–246 and references therein.
4. Davis, B.G. Synthesis of glycoproteins. Chem. Rev. 2002, 102, 579–601.
5. Kunz, H. Glycopeptides of biological interest: a challenge for chemical synthesis.
Pure Appl. Chem. 1993, 65, 1223–1232.
6. Wang, L. Chemoenzymatic synthesis of glycopeptides and glycoproteins through
endoglycosidase-catalyzed transglycosylation. Carbohydr. Res. 2008, 343, 1509–1522.
7. Anisfeld, S.T.; Lansbury, P.T. A convergent approach to the chemical synthesis of
asparagine-linked glycopeptides. J. Org. Chem. 1990, 55, 5560–5562.
8. Wang, Z.; Zhang, X.; Live, D.; Danishefsky, S.J. From glycols to glycopeptides: a
convergent and stereoselective total synthesis of a high mannose N-linked glycopeptide.
Angew. Chem. Int. Ed. 2000, 2000, 3652–3656.
9. Hojo, H.; Nakahara, Y. Recent progress in the solid-phase synthesis of glycopep-
tide. Curr. Protein Peptide Sci. 2000, 1, 23–48.
10. Arsequell, G.; Valencia, G. Recent advances in the synthesis of complex N-
glycopeptides. Tetrahedron Asymm. 1999, 10, 4045–3094.
11. Wen, S.; Guo, Z. Unprotected oligosaccharides as phase-tags: Solution-phase syn-
thesis of glycopeptides with solid-phase workups. Org. Lett. 2001, 3, 3773–3776.
12. Shao, N.; Xue, J.; Guo, Z. Chemical synthesis of CD52 glycopeptides containing
the acid-labile fucosyl linkage. J. Org. Chem. 2003, 68, 9003–9011.
13. Xue, J.; Guo, Z. Efficient synthesis of complex glycopeptides based on unprotected
oligosaccharides. J. Org. Chem. 2003, 68, 2713–2720.

Synthesis of N-Glycopeptides
113
14. Shao, N.; Guo, Z. Solution-phase synthesis with solid-state workup of an O-
glycopeptide with a cluster of cancer-related T antigens. Org. Lett. 2005, 7, 3589–3512.
15. Gololobov, Y.G.; Zhmurova, I.N.; Kasukhin, L.F. Sixty years of Staudinger reac-
tion. Tetrahedron 1981, 37, 437–472.
16. Koehn, M.; Breinbauer, R. The Staudinger ligation - A gift to chemical biology.
Angew. Chem. Int. Ed. 2004, 43, 3106–3116.
17. Hackenberger, C.P.R.; Schwarzer, D. Chemoselective ligation and modification
strategies for peptides and proteins. Angew. Chem. Int. Ed. 2008, 47, 10030–10074.
18. Nilsson, B.L.; Kiessling, L.L.; Raines, R.T. High-yielding Staudinger ligation of a
phosphinothioester and azide to form a peptide. Org. Lett. 2001, 3, 9–12.
19. Nilsson, B.L.; Hondal, R.J.; Soellner, M.B.; Raines, R.T. Protein assembly by or-
thogonal chemical ligation methods. J. Am. Chem. Soc. 2003, 125, 5268–5269.
20. He, Y.; Hinklin, R.J.; Chang, J.; Kiessling, L.L. Stereoselective N-glycosylation by
Staudinger ligation. Org. Lett. 2004, 6, 4479–4482.
21. Bianchi, A.; Bernardi, A. Traceless Staudinger ligation of glycosyl azides with tri-
aryl phosphines: stereoselective synthesis of glycosyl amides. J. Org. Chem. 2006, 71,
4565–4577.
22. Doores, K.J.; Mimura, Y.; Dwek, R.A.; Rudd, P.M.; Elliott, T.; Davis, B.G. Direct
deprotected glycosyl–asparagine ligation. Chem. Commun. 2006, 1401–1403.
23. Malkinson, J.P.; Falconer, R.A.; Toth, I. Synthesis of C-terminal glycopeptides from
resin-bound glycosyl azides via a modified Staudinger reaction. J. Org. Chem. 2000, 65,
5249–5252.
24. Schierholt, A.; Shaikh, H.A.; Schmidt-Lassen, J.; Lindhorst, T.K. Utilizing
Staudinger ligation for the synthesis of glycoamino acid building blocks and other gly-
comimetics. Eur. J. Org. Chem. 2009, 3783–3789.
25. Gyorgydeak, Z.; Szilagyi, L.; Paulsen, H. Synthesis, structure, and reactions of
glycosyl azides. J. Carbohydr. Chem. 1993, 12, 139–163.
26. Likhosherstov, L.M.; Novikova, O.S.; Derevitskaja, V.A.; Kochetkov, N.K. A new
simple synthesis of amino sugar β-D-glycosylamines. Carbohydr. Res. 1986, 146, C1–C5.
27. Vasella, A.; Witzig, C.; Chiara, J.L.; Martin-Lomas, M. Convenient synthesis of
2-azido-2-deoxy-aldoses by diazo transfer. Helv. Chim. Acta 1991, 74, 2073–2077.
28. Alper, P.B.; Hung, S.; Wong, C.-H. Metal catalyzed diazo transfer for the synthesis
of azides from amines. Tetrahedron Lett. 1996, 37, 6029–6032.
29. Dan, A.; Lergenmu¨ller, M.; Amano, M.; Nakahara, Y.; Ogawa, T.; Ito,
Y. p-Methoxybenzylidene-tethered β-mannosylation for atereoselective synthesis of
asparagine-linked glycan chains. Chem. Eur. J. 1998, 4, 2181–2190.
30. Shao, N.; Guo, Z. A facile synthesis of the fucosylated N-linked glycan core and
its application to solid-phase synthesis of CD52 glycopeptide. Pol. J. Chem. 2005, 79,
297–307.