Characterization of the action of tyrosinase on resorcinols.

The action of tyrosinase on resorcinol and some derivatives (4-ethylresorcinol, 2-methylresorcinol and 4-methylresorcinol) was investigated. If the catalytic cycle is completed with a reductant such as ascorbic acid or an o-diphenol such as 4-tert-butylcatechol, these compounds act as substrates of tyrosinase in all cases. The reaction can also be carried out, adding hydrogen peroxide to the medium. All the above compounds were characterized as substrates of the enzyme and their kinetic constants, KM (Michaelis constant) and kcat (catalytic constant) were determined. Measurement of the activity of the enzyme after pre-incubation with resorcinol, 4-ethylresorcinol or 4-methylresorcinol points to an apparent loss of activity at short times, which could correspond to an enzymatic inactivation process. However, if the measurements are extended over longer times, a burst is observed and the enzymatic activity is recovered, demonstrating that these compounds are not suicide substrates of the enzyme. These effects are not observed with 2-methylresorcinol. The docking results indicate that the binding of met-tyrosinase with these resorcinols occurs in the same way, but not with 2-methylresorcinol, due to steric hindrance.

Human insulin polymorphism upon ligand binding and pH variation: the case of 4-ethylresorcinol.

This study focuses on the effects of the organic ligand 4-ethylresorcinol on the crystal structure of human insulin using powder X-ray crystallography. For this purpose, systematic crystallization experiments have been conducted in the presence of the organic ligand and zinc ions within the pH range 4.50-8.20, while observing crystallization behaviour around the isoelectric point of insulin. High-throughput crystal screening was performed using a laboratory X-ray diffraction system. The most representative samples were selected for synchrotron X-ray diffraction measurements, which took place at the European Synchrotron Radiation Facility (ESRF) and the Swiss Light Source (SLS). Four different crystalline polymorphs have been identified. Among these, two new phases with monoclinic symmetry have been found, which are targets for the future development of microcrystalline insulin drugs.