Identification of two histidines as copper ligands in Streptomyces glaucescens tyrosinase

Abstract

The physiochemical properties of wild type and two mutants of Streptomyces glaucescens tyrosinase are reported. The native enzyme contains two coppers at the active site which are EPR nondetectable. The two coppers react stoichiometrically with one hydrogen peroxide molecule giving rise to oxytyrosinase. Its optical features are similar to those reported earlier for a molluscan hemocyanin. The two mutants in which histidine-62 and -189 were changed to asparagine by site-directed mutagenesis have lost their enzymatic activity and their ability to bind oxygen and contain only one copper ion which is fully EPR detectable. The EPR parameters indicate that the remaining copper is in a tetragonally distorted ligand environment. These data are in agreement with His-62 and His-189 serving as copper ligands in S. glaucescens tyrosinase.

Melanin production by Rhizobium meliloti GR4 is linked to nonsymbiotic plasmid pRmeGR4b: cloning, sequencing, and expression of the tyrosinase gene mepA

“…The conserved residues responsible for the copper binding activity (Fig. 5) seem to be present in the GR4 tyrosinase, although more detailed investigations will be necessary to prove whether the role of such residues is the same as that reported for other, homologous proteins of S. glaucescens (17), N. crassa (11), or 0. dofleini (22 (38). This activity for melanogenesis has been proposed to explain the survival of Vibrio cholerae in estuarine environments during the summer months, when water temperature rises and salinity increases because of evaporation (8).…”

“…3), and the CuB binding site is probably located between positions 223 and 267. A theoretical prediction of the potential role of these residues in binding copper is made possible by a comparison with similar residues that were previously proven to have this role (11,17,22). Thus, in the CuA binding site, the His-44 and His-53 positions are conserved; very likely the His-38 position is, too.…”

Identification of copper ligands in Aspergillus oryzae tyrosinase by site-directed mutagenesis

“…The EPR spectra of two mutants, His62Asn and His189Asn, of S. glaucescens were measured by Huber and Lerch [5]. The mutants had a more or less axially symmetrical appearance by these parameters.…”

“…Two histidine residues, His-62 and His-189, were identified as copper ligands in Streptomyces glaucescens tyrosinase by Huber and Lerch [5]. The native enzyme contains two copper atoms at the active site that are not detectable by EPR.…”

Transcription of the tyrosinase gene in Streptomyces michiganensis DSM 40015 is induced by copper and repressed by ammonium

“…In Streptomyces glaucescens the tyrosinase contains two copper atoms per molecule. Both copper atoms are bound to two histidine residues in the active centre of the enzyme and both are necessary for the activity of the enzyme (Huber & Lerch, 1988). In S. glaucescens, tyrosinase activity can be induced by various amino acids, including L-methionine, L-leucine and Lphenylalanine, but not L-tyrosine (Baumann & Kocher, 1976;Kieser et al, 1976), while the tyrosinase of Streptomyces antibioticus is induced at low effector concentrations only by L-methionine (Katz & Betancourt, 1988).…”