Albino mutants of Streptomyces glaucescens tyrosinase
| Autor: | Konrad Lerch, M P Jackman, A Hajnal |
|---|---|
| Rok vydání: | 1991 |
| Předmět: |
Albinism
Stereochemistry Tyrosinase Molecular Sequence Data Mutant Glycine Biochemistry Gene Expression Regulation Enzymologic Escherichia coli Histidine Amino Acid Sequence Site-directed mutagenesis Molecular Biology Base Sequence biology Monophenol Monooxygenase Mutagenesis Active site Gene Expression Regulation Bacterial Cell Biology biology.organism_classification Streptomyces Glutamine Genes Bacterial Luminescent Measurements Mutagenesis Site-Directed biology.protein Electrophoresis Polyacrylamide Gel Actinomycetales Research Article Chromatography Liquid Plasmids |
| Zdroj: | Biochemical Journal. 274:707-713 |
| ISSN: | 1470-8728 0264-6021 |
| DOI: | 10.1042/bj2740707 |
| Popis: | Site-directed mutagenesis was used to determine the functional role of several residues of Streptomyces glaucescens tyrosinase. Replacement of His-37, -53, -193 or -215 by glutamine yields albino phenotypes, as determined by expression on melanin-indicator plates. The purified mutant proteins display no detectable oxy-enzyme and increased Cu lability at the binuclear active site. The carbonyl derivatives of H189Q and H193Q luminesce, with lambda max. displaced more than 25 nm to a longer wavelength compared with native tyrosinase. The remaining histidine mutants display no detectable luminescence. The results are consistent with these histidine residues (together with His-62 and His-189 reported earlier) acting as Cu ligands in the Streptomyces glaucescens enzyme. Conservative substitution of the invariant Asn-190 by glutamine also gives an albino phenotype, no detectable oxy-enzyme and labilization of active-site Cu. The luminescence spectrum of carbonyl-N190Q, however, closely resembles that of the native enzyme under conditions promoting double Cu occupancy of the catalytic site. A critical role for Asn-190 in active-site hydrogen-bonding interactions is proposed. |
| Databáze: | OpenAIRE |
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