関: Edman sequencing

WordNet

changing to a lower state (a less respected state) (同)debasement

PrepTutorEJDIC

(地位の)格下げ / (品位などの)下落

Wikipedia preview

出典(authority):フリー百科事典『ウィキペディア（Wikipedia）』「2015/03/03 06:17:58」(JST)

wiki en

Edman degradation, developed by Pehr Edman, is a method of sequencing amino acids in a peptide.^[1] In this method, the amino-terminal residue is labeled and cleaved from the peptide without disrupting the peptide bonds between other amino acid residues.

Mechanism

Phenylisothiocyanate is reacted with an uncharged N-terminal amino group, under mildly alkaline conditions, to form a cyclical phenylthiocarbamoyl derivative. Then, under acidic conditions, this derivative of the terminal amino acid is cleaved as a thiazolinone derivative. The thiazolinone amino acid is then selectively extracted into an organic solvent and treated with acid to form the more stable phenylthiohydantoin (PTH)- amino acid derivative that can be identified by using chromatography or electrophoresis. This procedure can then be repeated again to identify the next amino acid. A major drawback to this technique is that the peptides being sequenced in this manner cannot have more than 50 to 60 residues (and in practice, under 30). The peptide length is limited due to the cyclical derivatization not always going to completion. The derivatization problem can be resolved by cleaving large peptides into smaller peptides before proceeding with the reaction. It is able to accurately sequence up to 30 amino acids with modern machines capable of over 99% efficiency per amino acid. An advantage of the Edman degradation is that it only uses 10 - 100 pico-moles of peptide for the sequencing process. The Edman degradation reaction was automated in 1967 by Edman and Beggs to speed up the process^[2] and 100 automated devices were in use worldwide by 1973.^[3]

Limitations

Because the Edman degradation proceeds from the N-terminus of the protein, it will not work if the N-terminal amino acid has been chemically modified or if it is concealed within the body of the protein. It also requires the use of either guesswork or a separate procedure to determine the positions of disulfide bridges, and peptide concentrations of 1 picomolar or above for discernible results.

Coupled analysis

Following 2D SDS PAGE the proteins can be transferred to a polyvinylidene difluoride (PVDF) blotting membrane for further analysis. Edman degradations can be performed directly from a PVDF membrane. N-terminal residue sequencing resulting in five to ten amino acid may be sufficient to identify a Protein of Interest (POI).

References

^ Edman, P.; Högfeldt, Erik; Sillén, Lars Gunnar; Kinell, Per-Olof (1950). "Method for determination of the amino acid sequence in peptides". Acta Chem. Scand. 4: 283–293. doi:10.3891/acta.chem.scand.04-0283 .
^ Edman P, Begg G (March 1967). "A protein sequenator". Eur J Biochem. 1 (1): 80–91. PMID 6059350.
^ Niall HD (1973). "Automated Edman degradation: the protein sequenator". Meth. Enzymol. Methods in Enzymology 27: 942–1010. doi:10.1016/S0076-6879(73)27039-8. ISBN 978-0-12-181890-6. PMID 4773306.

This chemical reaction article is a stub. You can help Wikipedia by expanding it.

UpToDate Contents

全文を閲覧するには購読必要です。 To read the full text you will need to subscribe.

1. 肝線維症の病因 pathogenesis of hepatic fibrosis
2. 肝線維症の新しい治療法 emerging therapies for hepatic fibrosis
3. 肝線維症の非侵襲的評価：血清学的検査および画像診断 noninvasive assessment of hepatic fibrosis overview of serologic and radiographic tests
4. 慢性腎疾患における炭水化物およびインスリン代謝 carbohydrate and insulin metabolism in chronic kidney disease
5. 副甲状腺ホルモンの分泌および作用 parathyroid hormone secretion and action

English Journal

Establishment of a mammalian expression system for recombinant [-2]proPSA and a specific antibody against the truncated leader peptide.

Hwang D1, Yoon A1, Kim S2, Kim H2, Chung J1,2.
Biotechnology and applied biochemistry.Biotechnol Appl Biochem.2016 Jan 21. doi: 10.1002/bab.1481. [Epub ahead of print]
A truncated precursor form of prostate-specific antigen (PSA), [-2]proPSA, is a well-known biomarker for prostate cancer. To develop a biomarker assay, highly purified [-2]proPSA is required as a standard reference and for generation of a specific antibody. In this study, we generated an efficient m
PMID 26790760

The Rise of Mass Spectrometry and the Fall of Edman Degradation.

Mann M1.
Clinical chemistry.Clin Chem.2016 Jan;62(1):293-4. doi: 10.1373/clinchem.2014.237271. Epub 2015 Oct 1.
PMID 26430071

Screening One-Bead-One-Compound Peptide Libraries for Optimal Kinase Substrates.

Trinh TB1, Pei D2.
Methods in molecular biology (Clifton, N.J.).Methods Mol Biol.2016;1360:169-81. doi: 10.1007/978-1-4939-3073-9_13.
Protein kinases phosphorylate specific serine, threonine, and/or tyrosine residues in their target proteins, resulting in functional changes of the target proteins such as enzymatic activity, cellular location, or association with other proteins. For many kinases, their in vivo substrate specificity
PMID 26501910

Japanese Journal

Isolation and Characterization of an Anti-Insect β-Toxin from the Venom of the Scorpion Isometrus maculatus

KAWACHI Tomoyuki,MIYASHITA Masahiro,NAKAGAWA Yoshiaki [他],MIYAGAWA Hisashi
Bioscience, biotechnology, and biochemistry 77(1), 205-207, 2013-01-23
Im-3 was isolated from the venom of the scorpion Isometrus maculatus through several steps of HPLC fractionation based on the insect paralytic activity. Injecting Im-3 into crickets induced paralysis, …
NAID 10031164715

リン酸化ペプチドの一次配列決定を可能にする次世代エドマン分解法

柴田孝之
旭硝子財団助成研究成果報告, 1-6, 2012
NAID 40019979786

〔報文〕シイタケ子実体のスーパーオキシドジスムターゼの精製と諸性質

不破眞佐子,池田啓一,川崎広明 [他],山倉文幸,高ひかり,峯木礼子,藤村務,松本孝
學苑 854, 1-9, 2011-12-01
… Direct analysis of N-terminal amino acid of this enzyme by Edman degradation using a protein sequencer cannot detect any amino acid residue, but amino acid residue of N-terminal appeared as serine residue after the treatment of the enzyme with tetrafluoroacetic acid in a vapor phase at 60℃ for 10min. Therefore, the N-terminal amino acid was modified with acetyl group. …
NAID 110008729425