Elucidating the structure of two cyclotides of Viola tianshanica maxim by MALDI TOF/TOF MS analysis / 药学学报
Acta Pharmaceutica Sinica
; (12): 1402-1409, 2010.
Article
in Zh
| WPRIM
| ID: wpr-353348
Responsible library:
WPRO
ABSTRACT
The cyclotides are a family of cyclic "mini" proteins that occur in Violaceae, Rubiaceae and Cucurbitaceae plant families and contain a head-to-tail cyclic backbone and a cystine knot arranged by three disulfide bonds. To study the natural cyclotides of V tianshanica, dried herb was extracted with 50% ethanol, and the concentrated aqueous extract was subjected to a solvent-solvent partitioning between water and hexane, ethyl acetate and n-butanol, separately. The n-butanol extract containing cyclotides was subjected to column chromatography over Sephadex LH-20, eluted with 30% methanol. The subfractions were directly reduced by DTT and analyzed by reverse-phase HPLC. The peaks with different retention times were shown on the profile of RP-HPLC and collected. The cyclotides were speculated based on masses range from 3 000 to 3 500 Da. The purified cyclotides were reduced with DTT, alkylated with iodoacetamide, and then were cleaved with endoproteinase Glu-C, endoproteinase Lys-C and Trypsin, separately. The digested peptides were purified on RP-HPLC and analyzed on MALDI TOF/TOF analyzer. A new cyclotide, cycloviolacin T1 and a reported cyclotide varv E were systemically determined using MALDI TOF/TOF system. So the method for the isolation and characterization of cyclotides was quickly built up in succession.
Full text:
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Index:
WPRIM
Main subject:
Plants, Medicinal
/
Molecular Sequence Data
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Molecular Structure
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Chemistry
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Chromatography, High Pressure Liquid
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Amino Acid Sequence
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Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
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Viola
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Cyclotides
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Tandem Mass Spectrometry
Language:
Zh
Journal:
Acta Pharmaceutica Sinica
Year:
2010
Type:
Article