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Glycoconjugates: synthesis and investigation of carbohydrate-protein interactions
Umeå University, Faculty of Science and Technology, Department of Chemistry. (Mikael Elofsson)
2010 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

To study the functions of glycoconjugates in biological systems reliable and efficient protocols for glycoconjugate synthesis are needed. To reach this goal we have developed methods for solid-phase synthesis of glycoconjugates that can be monitored with gel-phase 19F spectroscopy using fluorinated linkers, building blocks, and protecting groups. We have developed a new fluorine containing linker suitable for solid-phase synthesis of glycoconjugates. The linker was more acid-labile than similar linkers in order to enable cleavage under mild conditions of the target compound from the linker resin.  A carbamate-based strategy has been applied to attach a spacer carrying an amino group to a fluorinated Wang linker for synthesis of amino-functionalized glycoconjugates using thioglycoside donors with fluorinated protective groups. Cleavage from the solid support was performed with trifluoroacetic acid and subsequent protecting group removal gave the target compound. The terminal amine was conjugated with didecyl squarate and this derivative can be attached to various proteins and solid surfaces carrying primary or secondary amines. To evaluate this methodology we have immobilized glycoconjugates in amino-functionalized microtiter plates and successfully probed them with lectin. In addition, a novel fluorine containing protecting group has been designed, synthesized and evaluated. The protecting group was used for protection of the unreactive 4-OH in a galactose building block that was applied in the synthesis of 6-aminohexyl galabioside and was removed with TBAF in THF.

Adenovirus serotype 8 (Ad8), Ad19, and Ad37 cause the severe ocular infection, epidemic keratoconjunctivities (EKC). During infection, the adenoviruses interact with sialic acid containing glycoconjugates on the epithelial cells via fiber structures extending from the viral particles. The virus particle most likely binds to the host cell in a multivalent way by simultaneously using multiple fiber proteins and binding sites. Multivalent sialic acid containing conjugates could efficiently inhibit Ad37 cell attachment and subsequent infection of human corneal epithelial (HCE) cells. Three compact tri- and tetravalent sialic acid conjugates were prepared and evaluated as inhibitors of adenoviral host cell attachment and subsequent infection and all conjugates were potent as anti-adenoviral agents. The conjugates can readily be synthesized from accessible starting materials. A crystal structure of the Ad37 fiber knob protein and the trivalent sialic acid conjugate showed that the three binding sites were all occupied by one sialic acid residue each.

Place, publisher, year, edition, pages
Umeå: Kemiska institutionen, Umeå universitet , 2010. , 77 p.
Keyword [en]
Glycoconjugates, Carbohydrates, Galactose, Glucose, Solid-phase synthesis, SPOS, Glycosylation, Gel-phase 19F NMR spectroscopy, Fluorinated linker, Carbohydrate array, Microtiter plates, Carbohydrate-protein interactions, carbamate linker, Fsec, Protecting group, Fluorinated protecting group, Multivalent glycoconjugates, Adenovirus, Ad37, Epidemic keratoconjunctivitis, EKC, Sialic acid, Adenovirus inhibitor, Trivalent, Tetravalent, X-ray crystal structure
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
URN: urn:nbn:se:umu:diva-33841ISBN: 978-91-7459-014-2 (print)OAI: oai:DiVA.org:umu-33841DiVA: diva2:318255
Public defence
2010-06-04, KBC-huset, KB3B1, Umeå Universitet, Umeå, 10:00 (Swedish)
Opponent
Supervisors
Available from: 2010-05-14 Created: 2010-05-07 Last updated: 2010-05-19Bibliographically approved
List of papers
1. Synthesis and evaluation of 2-(2-fluoro-4-hydroxymethyl-5-methoxy-phenoxy)acetic acid as a linker in solid-phase synthesis monitored by gel-phase 19F NMR spectroscopy
Open this publication in new window or tab >>Synthesis and evaluation of 2-(2-fluoro-4-hydroxymethyl-5-methoxy-phenoxy)acetic acid as a linker in solid-phase synthesis monitored by gel-phase 19F NMR spectroscopy
2007 (English)In: Organic and biomolecular chemistry, ISSN 1477-0520, E-ISSN 1477-0539, Vol. 5, 2464-2471 p.Article in journal (Refereed) Published
Abstract [en]

Gel-phase 19F NMR spectroscopy is a useful monitoring technique for solid-phase organic chemistry due to the high information content it delivers and swift acquisition times, using standard NMR spectrometers. This paper describes the synthesis of the novel linker 2-(2-fluoro-4-hydroxymethyl-5-methoxy-phenoxy)acetic acid in 29% yield over seven steps, using nucleophilic aromatic substitutions on 2,4,5-trifluorobenzonitrile as key steps. Following standard solid-phase synthesis a peptide could be cleaved from the linker using 20% TFA in CH2Cl2 in 30 minutes, in contrast to a previously described monoalkoxy linker that requires 90% TFA in water at elevated temperature. A resin-bound peptide could be successfully glycosylated using only two equivalents of a thioglycoside donor, activated with N-iodosuccinimide and trifluoromethanesulfonic acid, and subsequent cleavage and deprotection gave the target glycopeptide. Direct glycosylation of the linker itself followed by mild acidic cleavage gave a fully protected hemiacetal for further chemical manipulation.

Place, publisher, year, edition, pages
The Royal Society of Chemistry, 2007
National Category
Other Basic Medicine
Identifiers
urn:nbn:se:umu:diva-15819 (URN)10.1039/b704472k (DOI)
Available from: 2007-08-02 Created: 2007-08-02 Last updated: 2010-05-12Bibliographically approved
2. Carbamate linker strategy in solid-phase synthesis of amino-functionalized glycoconjugates for attachment to solid surfaces and investigation of protein-carbohydrate interactions
Open this publication in new window or tab >>Carbamate linker strategy in solid-phase synthesis of amino-functionalized glycoconjugates for attachment to solid surfaces and investigation of protein-carbohydrate interactions
2009 (English)In: European Journal of Organic Chemistry, ISSN 1434-193X, E-ISSN 1099-0690, no 3, 349-57 p.Article in journal (Refereed) Published
Abstract [en]

Amino-functionalized serine-based galactose and glucose neoglycolipids were prepared by solid-phase synthesis using a carbamate strategy for anchoring amino functionalities to a (2-fluoro-4-hydroxymethylphenoxy)acetic acid linker resin. Key synthetic steps were monitored with gel-phase 19F NMR spectroscopy. Cleavage from the solid support was performed with trifluoroacetic acid. The terminal amine of the neoglycolipids was conjugated with didecyl squarate and then immobilized in amino-functionalized microtiter plates and the glycoconjugates were successfully probed with a galactose-binding lectin.

Place, publisher, year, edition, pages
Weinheim: WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim, 2009
Keyword
Solid-phase synthesis, Glycoconjugates, Carbohydrates, Proteins, Carbohydrate protein interactions, Carbamate linker, Gel-phase 19F NMR spectroscopy, Microtiter plates
National Category
Other Basic Medicine
Identifiers
urn:nbn:se:umu:diva-11489 (URN)10.1002/ejoc.200800670 (DOI)
Available from: 2009-01-12 Created: 2009-01-12 Last updated: 2010-05-12Bibliographically approved
3. Synthesis and application of a 2-[(4-fluorophenyl)sulfonyl]ethoxy carbonyl (Fsec) protected glycosyl donor in carbohydrate chemistry
Open this publication in new window or tab >>Synthesis and application of a 2-[(4-fluorophenyl)sulfonyl]ethoxy carbonyl (Fsec) protected glycosyl donor in carbohydrate chemistry
2010 (English)In: Molecules, ISSN 1420-3049, Vol. 15, no 8, 5708-5720 p.Article in journal (Refereed) Published
Abstract [en]

The 2-[(4-fluorophenyl)sulfonyl]ethoxy carbonyl (Fsec) group for protection of hydroxyl groups has been designed, synthesized, and evaluated. Fsec-Cl was readily prepared in 91% yield over three steps and subsequently used to protect 4-fluorobenzyl alcohol in high yield. The Fsec group was cleaved from the resulting model compound under mild basic conditions e.g., 20% piperidine in DMF and was stable under acidic conditions, e.g., neat acetic acid. The Fsec group was used to protect the unreactive 4-OH in a galactose building block that was later used in the synthesis of 6-aminohexyl galabioside.

Keyword
base sensitiveprotecting group, O-protection, glycoconjugate synthesis, glycosylation
National Category
Organic Chemistry
Research subject
Organic Chemistry
Identifiers
urn:nbn:se:umu:diva-33838 (URN)10.3390/molecules15085708 (DOI)000281481100039 ()
Available from: 2010-05-07 Created: 2010-05-07 Last updated: 2010-08-24Bibliographically approved
4. Synthesis and evaluation of tri- and tetravalent sialic acid inhibitors of EKC-causing adenoviruses
Open this publication in new window or tab >>Synthesis and evaluation of tri- and tetravalent sialic acid inhibitors of EKC-causing adenoviruses
Show others...
(English)Manuscript (preprint) (Other academic)
National Category
Microbiology in the medical area Organic Chemistry
Research subject
Organic Chemistry; Biorganic Chemistry; Microbiology
Identifiers
urn:nbn:se:umu:diva-33839 (URN)
Available from: 2010-05-07 Created: 2010-05-07 Last updated: 2010-05-12Bibliographically approved

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