- Mechanoenzymatic peptide and amide bond formation
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Mechanochemical chemoenzymatic peptide and amide bond formation catalysed by papain was studied by ball milling. Despite the high-energy mixing experienced inside the ball mill, the biocatalyst proved stable and highly efficient to catalyse the formation of α,α- and α,β-dipeptides. This strategy was further extended to the enzymatic acylation of amines by milling, and to the mechanosynthesis of a derivative of the valuable dipeptide L-alanyl-l-glutamine.
- Hernández, José G.,Ardila-Fierro, Karen J.,Crawford, Deborah,James, Stuart L.,Bolm, Carsten
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supporting information
p. 2620 - 2625
(2017/07/17)
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- Kinetically controlled peptide synthesis mediated by papain using the carbamoylmethyl ester as an acyl donor
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A series of dipeptides were synthesized generally in good yields with carbamoylmethyl (Cam) esters as acyl donors in the presence of a cysteine protease, papain, immobilized on Celite. Several segment condensations were also achieved generally in high yields without danger of racemization and formation of the secondary-hydrolysis product. Moreover, partial sequences of some bioactive peptides were prepared through segment condensations, and aimed-at peptides were obtained generally in high yields without the racemization of C-terminal residues of the carboxyl components. Thus, the superiority of the Cam ester in the kinetically controlled peptide synthesis was once again ascertained in couplings mediated by the cysteine protease as in those catalyzed by the serine proteases reported earlier.
- Miyazawa, Toshifumi,Horimoto, Takao,Tanaka, Kayoko
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p. 371 - 376
(2014/08/18)
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- Enzymatic C-terminal amidation of amino acids and peptides
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Herein, we describe two versatile and high yielding enzymatic approaches for the conversion of semi-protected amino acid and peptidyl C-terminal α-carboxylic acids into their corresponding amides. In the first approach, the lipase Candida antarctica lipase-B (Cal-B), and in the second approach, the protease Subtilisin A, are used, respectively. We found that by using the ammonium salt of the α-carboxylic acid instead of separate ammonia sources, the enzymatic amidation reactions proceeded much faster without side reactions and gave near to quantitative yields of products.
- Nuijens, Timo,Piva, Elena,Kruijtzer, John A.W.,Rijkers, Dirk T.S.,Liskamp, Rob M.J.,Quaedflieg, Peter J.L.M.
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experimental part
p. 3777 - 3779
(2012/09/22)
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- Fully enzymatic N→C-directed peptide synthesis using C-terminal peptide α-carboxamide to ester interconversion
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Chemoenzymatic peptide synthesis is potentially the most cost-efficient technology for the synthesis of short and medium-sized peptides with some important advantages. For instance, stoichiometric amounts of expensive coupling reagents are not required and racemisation does not occur rendering purification easier compared to chemical peptide synthesis. In this paper, a novel interconversion reaction of peptide C-terminal α-carboxamides into primary alkyl esters with alcalase was used to develop a fully enzymatic peptide synthesis strategy. For each elongation step a cost-efficient amino acid carboxamide building block was used followed by the interconversion of the elongated peptide carboxamide to the corresponding primary alkyl ester. These peptide esters are the starting materials for the next enzymatic peptide elongation step. Copyright
- Nuijens, Timo,Piva, Elena,Kruijtzer, John A. W.,Rijkers, Dirk T. S.,Liskamp, Rob M. J.,Quaedflieg, Peter J. L. M.
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experimental part
p. 1039 - 1044
(2011/07/09)
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- α-Chymotrypsin-catalyzed peptide synthesis in frozen aqueous solution using N-protected amino acid carbamoylmethyl esters as acyl donors
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A kinetically controlled peptide synthesis catalyzed by α-chymotrypsin was performed in frozen aqueous solution (ice, -24 °C). The yield of the peptide was significantly improved by the use of the carbamoylmethyl (Cam) ester as the acyl donor instead of the conventional ethyl ester. The peptide yield increased up to ca. 90% when N-benzyloxycarbonyl (CBZ)-Phe-OCam and H-Phe-NH2 were used as the acyl donor and nucleophile, respectively. Such an improvement of the peptide yield in ice was also observed in the coupling of other CBZ-amino acid Cam esters as acyl donors. Furthermore, this approach was applied to the synthesis of peptides containing d-amino acids. The peptides such as CBZ-d-Phe-Phe-NH2, CBZ-Phe-d-Phe-NH2 and CBZ-d-Phe-d-Phe-NH2 were also obtained in excellent to moderate yields in ice. A high diastereoselectivity towards the l-l peptide was observed when the racemic amino acid Cam ester was used as the acyl donor in ice.
- Salam, Sayed Mohiuddin Abdus,Kagawa, Ken-Ichi,Kawashiro, Katsuhiro
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- Reverse proteolysis promoted by in situ generated peptide ester fragments
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In this contribution we describe a general synthesis concept for the in situ preparation of protease specific reactants using methyl thioesters as universal precursors. The precursor esters are readily available by standard synthesis procedures and can be
- Wehofsky, Nicole,Koglin, Norman,Thust, Sven,Bordusa, Frank
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p. 6126 - 6133
(2007/10/03)
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- α-chymotrypsin-catalysed segment condensations via the kinetically controlled approach using carbamoylmethyl esters as acyl donors in organic media
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The superiority of the carbamoylmethyl ester as an acyl donor for the α-chymotrypsin-catalysed segment condensations via the kinetically controlled approach is demonstrated in several model systems carried out in organic media with low water content. Furthermore, this approach is successfully applied to the construction of the Leu-enkephalin sequence via a 4 + 1 segment coupling.
- Miyazawa, Toshifumi,Ensatsu, Eiichi,Hiramatsu, Makoto,Yanagihara, Ryoji,Yamada, Takashi
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p. 396 - 401
(2007/10/03)
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- α-chymotrypsin-catalysed peptide synthesis via the kinetically controlled approach using activated esters as acyl donors in organic solvents with low water content: Incorporation of non-protein amino acids into peptides
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The α-chymotrypsin-catalyzed peptide synthesis via the kinetically controlled approach using activated esters as acyl donors in orgnanic solvents with low water content was presented. The methyl esters of N-Z derivatives of racemic non-protein amino acids were chosen as carboxy components. They allowed the peptide-bond formation and optical resolution simultaneously to yield homochiral peptides. This method is useful for the incorporation of non-protein amino acids into peptides.
- Miyazawa, Toshifumi,Nakajo, Shin'ichi,Nishikawa, Miyako,Hamahara, Kazumi,Imagawa, Kiwamu,Ensatsu, Eiichi,Yanagihara, Ryoji,Yamada, Takashi
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- High isolated yields in thermodynamically controlled peptide synthesis in toluene catalysed by thermolysin adsorbed on Celite R-640
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An innovative immobilisation method that allows peptide synthesis to be performed even at equimolar concentrations, by controlling water activity, is reported.
- Basso, Alessandra,De Martin, Luigi,Ebert, Cynthia,Gardossi, Lucia,Linda, Paolo
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p. 467 - 468
(2007/10/03)
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- A novel support for enzyme adsorption: Properties and applications of aerogels in low water media
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Aerogels, because of their porosity, have a great ability to adsorb water, and this characteristic was exploited to extend the applicability of aerogels for the adsorption and entrapment of hydrolases to be used in organic media. The applicability of aero
- Basso,De Martin,Ebert,Gardossi,Tomat,Casarci,Li Rosi
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p. 8627 - 8630
(2007/10/03)
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- Superiority of the carbamoylmethyl ester as an acyl donor for the protease-catalyzed kinetically controlled peptide synthesis in organic media: Application to segment condensations
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The superiority of the carbamoylmethyl ester as an acyl donor for the α-chymotrypsin-catalyzed kinetically controlled peptide synthesis was demonstrated in several segment condensations carried out in organic media with low water content. Then this approach was successfully applied to the construction of the Leuenkephalin sequence via the 4 + 1 segment condensation.
- Miyazawa, Toshifumi,Ensatsu, Eiichi,Tanaka, Kayoko,Yanagihara, Ryoji,Yamada, Takashi
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p. 1013 - 1014
(2007/10/03)
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- Cross-linked crystals of subtilisin: Versatile catalyst for organic synthesis
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Cross-linked enzyme crystals (CLECs) of subtilisin exhibit excellent activity in aqueous and various organic solvents. This catalyst is more stable than the native enzyme in both aqueous and mixed aqueous/organic solutions. Subtilisin-CLEC was shown to be a versatile catalyst. It was used for the syntheses of peptides and peptidomimetics, mild hydrolysis of amino acid and peptide amides, enantio- and regioselective reactions, and transesterifications.
- Wang, Yi-Fong,Yakovlevsky, Kirill,Zhang, Bailing,Margolin, Alexey L.
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p. 3488 - 3495
(2007/10/03)
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- α-Chymotrypsin-catalysed peptide synthesis using activated esters as acyl donors
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The coupling efficiency in α-chymotrypsin-catalysed peptide synthesis is greatly improved by the use of activated esters such as the 2,2,2-trifluoroethyl ester as acyl donor instead of the conventional methyl ester; this approach is useful for the incorporation of non-protein amino acids into peptides.
- Miyazawa, Toshifumi,Nakajo, Shin'ichi,Nishikawa, Miyako,Imagawa, Kiwamu,Yanagihara, Ryoji,Yamada, Takashi
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p. 2867 - 2868
(2007/10/03)
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- Enzymatische Synthese von Peptiden und Ras-Lipopeptiden unter Verwendung des Cholinesters als loeslichkeitsvermittelnder Schutz- und Aktivierungsgruppe
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Keywords: Cholinesterase; Enzymkatalyse; Peptidsynthesen; Ras-Proteine
- Schelhaas, Michael,Glomsda, Simone,Haensler, Marion,Jakubke, Hans-Dieter,Waldmann, Herbert
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- ENZYMATIC PEPTIDE SYNTHESIS IN ORGANIC SOLVENT MEDIATED BY MODIFIED α-CHYMOTRYPSIN
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Peptide couplings have been catalyzed in organic medium containing a slight amount of water (0.5percent) by PEG modified α-Chymotrypsin.
- Babonneau, Marie-Therese,Jacquier, Robert,Lazaro, Rene,Viallefont, Philippe
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p. 2787 - 2790
(2007/10/02)
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- A Search for Peptide Ligase: Cosolvent-Mediated Conversion of Proteases to Esterases for Irreversible Synthesis of Peptides
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Serine and cysteine proteases (trypsin, chymotrypsin, papain, subtilisin) in the presence of certain concentrations of water-miscible organic solvents express no amidase activities.The esterase activities, however, remain significant.These modified enzyme
- Barbas, Carlos F. III,Matos, Jose R.,West, J. Blair,Wong, Chi-Huey
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p. 5162 - 5166
(2007/10/02)
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- Papain Catalysed Peptide Synthesis: Control of Amidase Activity and the Introduction of Unusual Amino Acids
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Procedures for the papain catalysed synthesis of peptides containing D-amino acids and derivatives with control of the enzyme's amidase activity have been developed.
- Barbas, Carlos F. III,Wong, Chi-Huey
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p. 533 - 534
(2007/10/02)
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- On the Use of Carboxamidomethyl Esters in the Protease-Catalyzed Peptide Synthesis
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Carboxyamidomethyl esters (CAM esters) of Z-and Boc-protected alanine and phenylalanine were prepared in order to investigate their usefulness as substrates for α-chymotrypsin- and papain-catalyzed hydrolysis and peptide synthesis reactions.The easy removal of the CAM-C-protecting group under mild conditions and dependent on the enzyme specificity was demonstrated.Examples are given for the protease-catalyzed synthesis of various peptide derivatives using CAM esters as C- and N-components in aqueous-organic media.Comparatively short reaction times were observed. - Key words: Carboxyamidomethyl ester as C-protecting group; Enzymatic deprotection; Peptide synthesis; α-Chymotrypsin- and Papain-catalyzed peptide bond formation
- Kuhl, Peter,Zacharias, Ute,Burckhardt, Helmut,Jakubke, Hans-Dieter
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p. 1195 - 1204
(2007/10/02)
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- Solubilizing Protective Groups for Enzymatic Peptide Syntheses. Studies with Polyoxyethylene-Supported Substrates
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The utility of amino acids and dipeptides esterified to solubilizing polyoxyethylene supports as substrates for protease-mediated peptide bond formation was studied using free and immobilized α-chymotrypsin as a catalyst.Poor yields were obtained with Nα-protected amino acid or dipeptide polyoxyethylene esters as carboxyl components, most probably due to a shielding of the active site of the acyl enzyme intermediate by the polymer favouring hydrolytic cleavage.Experiments with polyoxyethylene esters as nucleophiles gave good results with free chymotrypsin, immobilized chymotrypsin predominantly caused hydrolysis of the ester carboxyl component due to unfavourable interactions between the soluble and insoluble polymers.Keywords: Chymotrypsin; Enzymatic peptide synthesis; Liquid phase peptide synthesis; Solubilizing protective groups; Polyoxyethylene
- Koennecke, Andreas,Pchalek, Volker,Jakubke, Hans-Dieter
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p. 111 - 118
(2007/10/02)
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- Peptide Synthesis by Means of Immobilized Enzymes II. Immobilized Trypsin, Thermolysin and Papain
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Model studies were performed on the utility of covalently immobilized trypsin, thermolysin and papain for peptide bond formation.Trypsin and thermolysin catalyzed the formation of peptide bonds with nearly the same efficiency as the soluble proteases and they could be re-used successfully for further coupling experiments.The possibility of using immobilized trypsin and papain for kinetically controlled peptide bond formation was investigated.With the serine type enzyme trypsin excellent product yields were obtained starting with ester carboxyl components and an economical ratio of substrates.Experiments with the thiol protease papain were unsatisfactory because the once formed product is hydrolyzed as fact as the starting ester substrate used. - Keywords: Immobilized enzymes; Papain; Peptide synthesis; Thermolysin; Trypsin
- Koennecke, Andreas,Haensler, Marion,Schellenberger, Volker,Jakubke, Hans-Dieter
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p. 433 - 444
(2007/10/02)
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- Catalytic Transfer Hydrogenation in Pepetide Synthesis: Synthesis of 5- and 5-enkephalins
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The simplicity in the removal of N-protecting groups like benzyloxycarbonyl employed in peptide synthesis by catalytic transfer hydrogenation at room temperature using formic acid in presence of palladium black has been demonstrated by the synthesis of the opioid pentapeptides 5- and 5-enkephalins.
- Sivanandaiah, K. M.,Gurusiddappa, S.
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p. 857 - 859
(2007/10/02)
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- Synthesis of Amides of Dipeptides and Kinetics of Papain-catalysed Hydrolysis of These Amides
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The N-benzyloxycarbonylated amides of the dipeptides L-alanyl-L-phenylalanine, L-leucylleucine, L-leucyl-L-phenylalanine, L-phenylalanyl-L-alanine and L-phenylalanyl-L-leucine have been synthesised and the kinetic parameters for the papain-catalysed hydrolysis of these substrates and their ester analogs have been determined at pH 8 and 9.All the amide substrates are hydrolysed at the amide linkage except N-benzyloxycarbonyl-L-alanyl-L-phenylalanine amide which hydrolyses to the extent of 45percent at the amide linkage and 55percent at the peptide linkage.For amide hydrolysis acetylation and deacetylation rate constants are approximately equal.
- Zaher, M. R.,El-Sharief, A. M. Sh.
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p. 740 - 743
(2007/10/02)
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