Homogenization and crystallization of histidine ammonia-lyase by exchange of a surface cysteine residue |
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| Authors: | Schwede, Torsten F. Badeker, Mathias Langer, Martin Retey, Janos Schulz, Georg E. |
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| Affiliation: | 1 Institut für Organische Chemie und Biochemie, Albertstrasse 21,D-79104 Freiburg im Breisgau and 2 Institut für Organische Chemie, Lehrstuhl Biochemie der Universität, Richard-Willstätter-Allee, D-76128 Karlsruhe, Germany |
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| Abstract: | Histidase (histidine ammonia-lyase, EC 4.3.1.3) from Pseudomonasputida was expressed in Escherichia coli and purified. In theabsence of thiols the tetrameric enzyme gave rise to undefinedaggregates and suitable crystals could not be obtained. Thesolvent accessibility along the chain was predicted from theamino acid sequence. Among the seven cysteines, only one waslabeled as `solvent-exposed'. The exchange of this cysteineto alanine abolished all undefined aggregations and yieldedreadily crystals diffracting to 1.8 Å resolution. |
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| Keywords: | disulfide engineering/ Ellman's reaction/ histidine ammonia-lyase/ unspecific aggregation/ X-ray crystallography |
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