DOI: 10.1002/bit.260261003. Peptide synthesis in aqueous–organic solvent mixtures with α‐chymotrypsin immobilized to tresyl chloride‐activated agarose

Peptide synthesis in aqueous–organic solvent mixtures with α‐chymotrypsin immobilized to tresyl chloride‐activated agarose

10.1002/bit.260261003

Crossref journal-article

Wiley

Biotechnology and Bioengineering (311)

Abstract

Abstractα–Chymotrypsin was immobilized with a high coupling yield (up to 80%) to tresyl chloride activated Sepharose CL‐4B.The immobilized enzyme was tested for its ability to synthesize soluble peptides from N‐acetylated amino acid esters as acyl donors and amino acid amides as acceptor amines in water–water‐miscible organic solvent mixtures. It was found that the yield of peptide increased with increasing concentration of organic cosolvent. Almost complete synthesis (97%) of Ac–Phe–Ala–NH2 was obtained from Ac–Phe–OMe using a sixfold excess of Ala–NH2. The rate of peptide formation in aqueous–organic solvent mixtures was good. Thus, 0.1M peptide was formed in less than 2 h in 50 vol% DMF with 0.1 mg immobilized chymotrypsin/mL reaction mixture. The immobilized enzyme distinguished between the L and D configurations of acceptor amino acid amides even in high concentration of nonaqueous component (90% 1,4‐butanediol). The effect of temperature was studied. It was found that both the yield of peptide and the stability of immobilized enzyme increased when the temperature was lowered. Experiments could be performed at subzero temperatures in the aqueous–organic solvent mixtures resulting in very high yield of peptide. After three weeks continuous operation at 4°C in 50% DMF, the immobilized enzyme retained 66%of its original synthetic activity. The activity of the immobilized enzyme was better conserved with a preparation made from agarose with a higher tresyl group content compared to a preparation made from a lower activated agarose, indicating that multiple point of attachment has a favorable effect on the stability of the enzyme in aqueous–organic solvent mixtures. The major advantage of using water‐miscible instead of water‐immiscible organic solvents to promote peptide syntheses appears to be the increased solubility of substrates and products, making continuous operation possible.

Bibliography

Nilsson, K., & Mosbach, K. (1984). Peptide synthesis in aqueousâorganic solvent mixtures with Î±âchymotrypsin immobilized to tresyl chlorideâactivated agarose. Biotechnology and Bioengineering, 26(10), 1146â1154. Portico.

Authors 2

Kurt Nilsson (first)
Klaus Mosbach (additional)

References 34 Referenced 83

10.1016/S0021-9258(18)74417-7 / J. Biol. Chem. by Bergmann M. (1937)
10.1002/9780470122983.ch7
10.1007/BF02798320
10.1007/BF02910870
10.1016/S0040-4039(00)77731-8
10.1016/0005-2744(81)90252-7
10.1021/jo00338a045
10.1021/bi00629a009
{'key': 'e_1_2_1_10_2', 'first-page': '157', 'volume': '45', 'author': 'Douzo P.', 'year': '1977', 'journal-title': 'Adv. Enzymol.'} / Adv. Enzymol. by Douzo P. (1977)
10.1002/9780470122976.ch3
10.1016/0006-291X(81)91541-2
{'key': 'e_1_2_1_13_2', 'volume-title': 'Chemistry of the Amino Acids', 'author': 'Greenstein J. P.', 'year': '1961'} / Chemistry of the Amino Acids by Greenstein J. P. (1961)
10.1016/S0003-9861(57)80011-3
10.1007/978-3-642-86602-9
{'key': 'e_1_2_1_16_2', 'first-page': '56', 'volume-title': 'Methods in Enzymology', 'author': 'Nilsson K.', 'year': '1984'} / Methods in Enzymology by Nilsson K. (1984)
10.1021/bi00735a001
10.1042/bj1630531
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{'key': 'e_1_2_1_21_2', 'volume-title': 'Enzyme Structure and Mechanism', 'author': 'Fersht A.', 'year': '1977'} / Enzyme Structure and Mechanism by Fersht A. (1977)
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10.1111/j.1432-1033.1973.tb02689.x
10.1016/0021-9797(76)90076-X
10.1021/bi00519a023
10.1021/ja01009a037
T.Mori K.Nilsson P.‐O.Larsson andK.Mosbach submitted for publication.
10.1016/S0021-9258(19)70636-X / J. Biol. Chem. by Kullman W. (1980)

Dates

Type	When
Created	20 years, 8 months ago (Dec. 29, 2004, 10:49 p.m.)
Deposited	1 year, 10 months ago (Oct. 19, 2023, 5:15 p.m.)
Indexed	1 day, 19 hours ago (Sept. 4, 2025, 10:34 a.m.)
Issued	40 years, 11 months ago (Oct. 1, 1984)
Published	40 years, 11 months ago (Oct. 1, 1984)
Published Online	21 years, 6 months ago (Feb. 18, 2004)
Published Print	40 years, 11 months ago (Oct. 1, 1984)

Funders 0

None

BibTeX

@article{Nilsson_1984, title={Peptide synthesis in aqueous–organic solvent mixtures with α‐chymotrypsin immobilized to tresyl chloride‐activated agarose}, volume={26}, ISSN={1097-0290}, url={http://dx.doi.org/10.1002/bit.260261003}, DOI={10.1002/bit.260261003}, number={10}, journal={Biotechnology and Bioengineering}, publisher={Wiley}, author={Nilsson, Kurt and Mosbach, Klaus}, year={1984}, month=oct, pages={1146–1154} }

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