| Project/Area Number |
60870076
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| Research Category |
Grant-in-Aid for Developmental Scientific Research
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| Allocation Type | Single-year Grants |
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| Research Field |
Chemical pharmacy
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| Research Institution | Kyoto University |
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Principal Investigator |
YAJIMA Haruaki Faculty of pharmaceutical Sciences, Kyoto University, 薬学部, 教授 (00025678)
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| Co-Investigator(Kenkyū-buntansha) |
AKAJI Kenichi Faculty of Pharmaceutical Sciences, Kyoto University, 薬学部, 助手 (60142296)
FUNAKOSHI Susumu Faculty of Pharmaceutical Sciences, Kyoto University, 薬学部, 助手 (10135593)
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| Project Period (FY) |
1985 – 1987
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| Project Status |
Completed (Fiscal Year 1987)
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| Budget Amount *help |
¥5,900,000 (Direct Cost: ¥5,900,000)
Fiscal Year 1987: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1986: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1985: ¥3,900,000 (Direct Cost: ¥3,900,000)
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| Keywords | Human insulin-like growth factor-I / Trimethylsilyl trifluoromethanesulfonate / トリメチルシリルブロマイド / インスリン様成長因子 / 1-アダマンチルシステイン / トリメチルシリルトリフルオロメタンスルホネート脱保護法 / β-シクロヘプチルアスパルテイト / トリフルオロメタン脱保護法 |
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| Research Abstract |
In 1978, Rinderknecht and Humbel elucidated the complete amino acid sequence of human insulin-like growh factor I (hiGF-I) and this structure was later confirmes by cDNA sequence analysis of its precursor. Because of grest interests in physiological roles of this peinciple associated with growht hormone, we undertook the synthesis of a 70-residue peptide corresponding to the entire amino acid sequence of hIGF-I with three disulfide bridges. First we prepared a linear form of protected hIGF-I by assembling 13-peptide fragments of established purity. Final deprotection and disulfide bond formation were conducted and the product was purified to homogeneity by HPLC, as outlined below.
Prior to deprotection, Met(O) was reduced back to Met by treatment with phenyl thiotrimethylsilane. To ensure complete removal of all protecting groups attached, deprotection was carried out in 2 steps: 1) Removal of Bzl-type protecting groups by lM trimethylsilyl bromide-thioanisole/TFS + EDT at 0゜C for 3h. 2) Removal of other protecting groups by 1M trimethylsilyl trifuloromethanesulfonate-thioanisole/TFA + EDT at 0゜C for 3h. The deprotected peptide was air-oxidized in the presence of reduced and oxidized glutathione. After ion-exchange chromatography, followed by HPLC purification, the product possesing an identical HPLC retention time with that of biosynthetic hIGF-I was obtained (yield 0.79%, from protected peptide). Improvement of the yield and biological evaluation are under inverstigation.
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