Human alcohol dehydrogenase (ADH, tiff isozyme of class I) was expressed in Escherichia coli, purified to homogeneity, and characterized regarding N-terminal processing. The expression system was obtained by ligation of a cDNA fragment corresponding to the fl-subunit of human liver alcohol dehydrogenase into the vector pKK 223-3 containing the tac promoter. The enzyme, detected by Western-blot analysis and ethanol oxidizing activity, constituted up to 3% of the total amount of protein. Recombinant ADH was separated from E. coli ADH by ion-exchange chromatography and the isolated enzyme was essentially pure as judged by SDS-polyacrylamide gel electrophoresis and sequence analysis. The N-terminal sequence was identical to that of the authentic fl-subunit except that the N-terminus was non-acetylated, indicating a correct removal of the initiator methionine, but lack of further processing.
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December 01 1987
Expression in Escherichia coli of active human alcohol dehydrogenase lacking N-terminal acetylation
Jan-Olov Höög;
Jan-Olov Höög
1Department of Chemistry I, Karolinska Institutet, S-104 01 Stockholm, Sweden
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Marianne Weis;
Marianne Weis
1Department of Chemistry I, Karolinska Institutet, S-104 01 Stockholm, Sweden
2Department of Biochemistry, Universität des Saarlandes, D-6600 Saarbrücken, FRG
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Michael Zeppezauer;
Michael Zeppezauer
2Department of Biochemistry, Universität des Saarlandes, D-6600 Saarbrücken, FRG
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Hans Jörnvall;
Hans Jörnvall
1Department of Chemistry I, Karolinska Institutet, S-104 01 Stockholm, Sweden
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Hedvig von Bahr-Lindstrom
Hedvig von Bahr-Lindstrom
1Department of Chemistry I, Karolinska Institutet, S-104 01 Stockholm, Sweden
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Publisher: Portland Press Ltd
Received:
January 15 1988
Online ISSN: 1573-4935
Print ISSN: 0144-8463
© 1987 Plenum Publishing Corporation
1987
Biosci Rep (1987) 7 (12): 969–974.
Article history
Received:
January 15 1988
Citation
Jan-Olov Höög, Marianne Weis, Michael Zeppezauer, Hans Jörnvall, Hedvig von Bahr-Lindstrom; Expression in Escherichia coli of active human alcohol dehydrogenase lacking N-terminal acetylation. Biosci Rep 1 December 1987; 7 (12): 969–974. doi: https://doi.org/10.1007/BF01122131
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