NDLI: High-level expression and deletion mutagenesis of human tryptophan hydroxylase.

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High-level expression and deletion mutagenesis of human tryptophan hydroxylase.

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High-level expression and deletion mutagenesis of human tryptophan hydroxylase.

Content Provider	World Health Organization (WHO)-Global Index Medicus
Author	Yang, X. J. Kaufman, S.
Description	Author Affiliation: Yang XJ ( Laboratory of Neurochemistry, National Institute of Mental Health, National Institutes of Health, Bethesda, MD 20892.);
Abstract	Human tryptophan hydroxylase has been expressed as a soluble and active form in Escherichia coli by fusion with an affinity tag, maltose-binding protein. The fusion protein has been purified to near homogeneity by affinity chromatography on crosslinked amylose resin. The purified fusion protein has a specific activity of 86 nmol of 5-hydroxytryptophan per min per mg of fusion protein. A series of truncation mutants have also been made to explore the domain organization of tryptophan hydroxylase. All deletion mutants were subject to affinity purification and kinetic characterization. While removal of the N-terminal 164 amino acids completely inactivates the enzyme, deletion of the first 91 residues results in a 7-fold reduction in specific activity. From the C terminus, deletion of 36, 55, or 112 amino acids abolishes the activity, whereas deletion of 19 residues decreases the specific activity by approximately 11-fold. These results are consistent with a model for tryptophan hydroxylase in which the enzyme consists of an N-terminal regulatory domain, a catalytic core, and a small C-terminal region of uncertain but important function.
ISSN	00278424
e-ISSN	10916490
Journal	Proceedings of the National Academy of Sciences of the United States of America
Issue Number	14
Volume Number	91
Language	English
Publisher	National Academy of Sciences
Publisher Date	1994-08-01
Publisher Place	United States
Access Restriction	Open
Subject Keyword	ATP-Binding Cassette Transporters Escherichia Coli Proteins Monosaccharide Transport Proteins Mutagenesis, Site-Directed Sequence Deletion Tryptophan Hydroxylase Metabolism Blotting, Western Carrier Proteins Biosynthesis Isolation & Purification DNA Primers Escherichia Coli Genetics Genetic Vectors Kinetics Maltose-Binding Proteins Molecular Sequence Data Plasmids Recombinant Fusion Proteins Restriction Mapping Comparative Study Multidisciplinary
Content Type	Text
Resource Type	Article
Subject	Multidisciplinary

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