Silver trifluoromethanesulphonate as an S-deprotecting reagent for the synthesis of cystine peptides

Fujii, Nobutaka; Otaka, Akira; Watanabe, Toshihiro; Okamachi, Akira; Tamamura, Hirokazu; Yajima, Haruaki; Inagaki, Yoshimasa; Nomizu, Motoyoshi; Asano, Kimiko

doi:10.1039/c39890000283

Cited by 58 publications

(42 citation statements)

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“…However MALDI-MS analysis indicated that decomposition of the formed Hg-thiolate compounds turned out to be more challenging and required longer reaction times and a larger excess of DTT when compared with the corresponding Ag-thiolates (data not shown). For this reason, we chose to stick to the AgOAc protocol, which also has been reported to be milder in the presence of sensitive amino acid residues (31,32).…”

Section: Resultsmentioning

confidence: 99%

Convergent chemical synthesis and high-resolution x-ray structure of human lysozyme

Durek

Torbeev

Kent

2007

Proc. Natl. Acad. Sci. U.S.A.

160

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In this article, we report the total chemical synthesis of human lysozyme. Lysozyme serves as a widespread model system in various fields of biochemical research, including protein folding, enzyme catalysis, and amyloidogenesis. The 130-aa wild-type polypeptide chain of the human enzyme was assembled from four polypeptide segments by using native chemical ligation in a fully convergent fashion. Key to the assembly strategy is the application of the recently developed kinetically controlled ligation methodology, which provides efficient control over the ligation of two peptide ␣ thioesters to yield a unique product. This result enables the facile preparation of a 64-residue peptide ␣ thioester; this segment is joined by native chemical ligation to a 66-aa Cys peptide, to yield the target 130-aa polypeptide chain. The synthetic polypeptide chain was folded in vitro into a defined tertiary structure with concomitant formation of four disulfides, as shown by 2D TOCSY NMR spectroscopy. The structure of the synthetic human lysozyme was confirmed by high-resolution x-ray diffraction, giving the highest-resolution structure (1.04 Å) observed to date for this enzyme. Synthetic lysozyme was obtained in good yield and excellent purity and had full enzymatic activity. This facile and efficient convergent synthesis scheme will enable preparation of unique chemical analogs of the lysozyme molecule and will prove useful in numerous areas of lysozyme research in the future. Lysozyme is possibly one of the best studied enzymes. The x-ray structure of hen egg-white lysozyme, initially reported in 1965, was the first high-resolution 3D structure of an enzyme molecule (1). Since then the protein has served as a model system for the study of protein folding and misfolding, enzyme catalysis and mechanism, x-ray crystallography, enzyme evolution, and protein engineering (2-9). Moreover, human lysozyme recently has attracted considerable interest because certain mutations in the enzyme were shown to render the protein amyloidogenic (10, 11).Despite extensive genetic, structural, and physico-chemical studies carried out over the last 50 years, many questions regarding lysozyme folding, catalysis, and amyloid fibril formation remain unsolved, unsatisfactorily explained, or controversial. This deficit is at least in part attributable to the limited means that could be used to modify the chemical structure of the lysozyme molecule. More powerful and versatile control over the structure of the enzyme is required for a detailed understanding of the properties of the protein on the molecular or atomic scale. Chemical protein synthesis has emerged as a powerful tool in this respect, especially because it grants nearly absolute control over the covalent structure of an enzyme molecule (12)(13)(14). Given the widespread and longlasting interest in lysozyme research, it is not surprising that for exactly these reasons chemical synthesis of the full-length protein has been envisioned (and attempted) as early as the 1970s (15, 16). However, these ...

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Section: Resultsmentioning

confidence: 99%

Convergent chemical synthesis and high-resolution x-ray structure of human lysozyme

Durek

Torbeev

Kent

2007

Proc. Natl. Acad. Sci. U.S.A.

160

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“…The crude reaction mixture was directly subjected to MFD. Happily, Cys 34 was smoothly reduced to Ala 34 (3→4) whereas the Thr 54 -SEt functional group remained intact. NCL of fragments 3a (6) and 4a (7) followed by treatment with MeONH 2 •HCl provided polypeptide 8 with all of the cysteine residues deprotected.…”

Section: Synthesis Of the Gm-csf Aglyconementioning

confidence: 99%

Synthesis of granulocyte–macrophage colony-stimulating factor as homogeneous glycoforms and early comparisons with yeast cell-derived material

Zhang

Johnston

Shieh

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

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Significance As biologically active glycoproteins are increasingly investigated as potential therapeutic agents, there is a growing demand for the development of strategies for the synthesis of homogeneous, single-glycoform constructs for the purposes of rigorous evaluation. Currently, most glycoproteins are accessed through recombinant methods as complex mixtures of glycoforms. Granulocyte–macrophage colony-stimulating factor (GM-CSF) is an important glycoprotein therapeutic, used to stimulate the immune system following bone-marrow transplant and chemotherapy. GM-CSF is also being explored as a potential immunoadjuvant for anticancer vaccines. We have completed a chemical synthesis of homogeneous, single-glycoform GM-CSF. Through adaptation of this modular synthetic route, it will be possible to gain access to a menu of single-glycoform GM-CSF congeners for a wide range of biological studies.

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“…They are also quite stable to HF at low temperatures but cleaved at higher temperatures in the presence of scavengers. 335 They are also stable to Ag (I) trifluoromethanesulfonate in TFA, 344 which quantitatively removes the S-Mmt group, and also to iodine oxidation. Other possible cleavage conditions are listed in the table.…”

Section: Protecting Groups Removed By Acidmentioning

confidence: 99%

“…343 It can be selectively removed in the presence of Meb using Ag(I) trifluoromethanesulfonate in TFA. 344 An intramolecular disulfide bridge between two Cys(Mob)-protected residues can be formed by removing the Mob group with MeSiCl 3 or SiCl 4 in TFA in the presence of diphenyl sulfoxide at 4ºC in 30 min. 345 In addition, oxidative removal with Tl(III) trifluoroacetate also leads to the formation of a disulfide bridge by reaction with a free Cys side chain.…”

Section: Protecting Groups Removed By Acidmentioning

confidence: 99%

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Amino Acid-Protecting Groups

2009

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Silver trifluoromethanesulphonate as an S-deprotecting reagent for the synthesis of cystine peptides

Cited by 58 publications

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Convergent chemical synthesis and high-resolution x-ray structure of human lysozyme

Convergent chemical synthesis and high-resolution x-ray structure of human lysozyme

Synthesis of granulocyte–macrophage colony-stimulating factor as homogeneous glycoforms and early comparisons with yeast cell-derived material

Amino Acid-Protecting Groups

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