Precise chemical synthesis of glycorpoteins for their structural analysis by nuclear magnetic resonance spectroscopy
| Project/Area Number |
15K13742
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| Research Category |
Grant-in-Aid for Challenging Exploratory Research
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| Allocation Type | Multi-year Fund |
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| Research Field |
Bio-related chemistry
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| Research Institution | Osaka University |
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Principal Investigator |
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| Project Period (FY) |
2015-04-01 – 2017-03-31
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| Project Status |
Completed (Fiscal Year 2016)
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| Budget Amount *help |
¥4,030,000 (Direct Cost: ¥3,100,000、Indirect Cost: ¥930,000)
Fiscal Year 2016: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2015: ¥2,080,000 (Direct Cost: ¥1,600,000、Indirect Cost: ¥480,000)
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| Keywords | 糖質関連化学 / 糖鎖工学 / 糖タンパク質 / 糖鎖 / 核磁気共鳴法 / 安定同位体 / 安定同位体標識 |
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| Outline of Final Research Achievements |
Using solid-phase peptide synthesis (SPPS) and native chemical ligation (NCL), thirteen 15N-labeled amino acids were inserted at specific positions of the protein backbone, while intentionally varying the enrichment of 15N atoms. This method enabled us to assign 1H-15N signals of synthetic glycoproteins even the same type of amino acid based on the intensities of 1H-15N HSQC signals. We synthesized three kinds of glycoproteins bearing an oligosaccharide of either a bi-antennary complex-type or a high-mannose-type and non-glycosylated one. Our NMR experiments showed that glycosylation at the native glycosylation positions did not disturb protein conformation. However, temperature-varied circular dichroism (CD) spectra and T1 values indicated that oligosaccharides appeared to inhibit protein fluctuation.
We also demonstrated novel semisynthesis of tribrached complex type oligosaccharide which can prepared within 10 conversion steps from the isolated biantennary oligosaccharide.
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Report
(3 results)
Research Products
(5 results)
