Evaluation of nano-sized virgin coconut oil (VCO)-loaded liposomes for enhancing mushroom and B16-F10 tyrosinase activity

Non-specific binding sites help to explain mixed inhibition in mushroom tyrosinase activities

Inhibitory effects of cinnamic acid and its derivatives on the diphenolase activity of mushroom ( Agaricus bisporus) tyrosinase

Objective To study the effects of N-acetylglucosamine on the activity of mushroom tyrosinase in vitro as well as on tyrosinase activity and melanogenesis in B16 melanoma cells. Methods Various concentrations (1.5 - 50 mmol/L) of N-acetylglucosamine were used to incubate with mushroom tyrosinase for 10 minutes following by the measurement of dopa oxidase activity of tyrosinase. B16 melanoma cells were cultured and treated with six concentrations (1.5 - 100 mmol/L) of N-acetylglu-cosamine for 72 hours; then, the cell proliferation of, melanogenesis and expression of tyrosinase protein and activity of tyrosinase in cultured B16 cells were measured by methyl thiazolyl tetrazolium (MTT) assay, sodium hydroxide (NaOH)-lysis method, immunocytochemical method and dopa oxidation assay, respectively. Results N-acetylglucosamine of 1.5, 3, 6.25, 25 and 50 mmol/L significantly inhibited the activity of mush-room tyrosinase compared with phosphate buffered saline (0.7380 ± 0.0254, 0.7293 ± 0.0382, 0.7247 ± 0.0389, 0.7233 ± 0.0186, 0.7043 ± 0.0166 vs 0.8183 ± 0.0326, P < 0.05 or 0.01). After treatment with N-acetylglucosamine of 25, 50 and 100 mmol/L, a significant suppression was observed in cell proliferation (ahsorbance: 0.5410 ± 0.0496, 0.4480 ± 0.0246 and 0.1273 ± 0.0137 vs 0.6523 ± 0.0569) of and melano-genesis (absorbance at 460 nm: 0.0070 ± 0.0008, 0.0049 ± 0.0012 and 0.0015 ± 0.0014 vs 0.0096 ± 0.0014) in B16 melanoma cells. Also, decreased activity (absorbance at 460 nm: 0.1003 ± 0.0404 and 0.0130 ± 0.0053 vs 0.2283 ± 0.0691 ) and protein expression ( 13.2700 ± 0.9741 and 8.5667 ± 2.0345 vs 17.4703 ± 2.0583 ) of tyrosinase were noted in B16 cells treated with N-acetylglucosamine of 50 and 100 mmol/L. Conclusions These studies show that N-acetylglucosamine inhibits tyrosinase activity and melanogenesis in murine B16 melanoma cells. Hence, N-acetylglucosamine may serve as a skin lightening agent in the future. Key words: Tumor cells; cultured; Monophenol monooxygenase; Melanins; N-acetylglucosamine

Catecholase and cresolase activities of mushroom tyrosinase (MT) were studied in presence of some n-alkyl carboxylic acid derivatives. Catecholase activity of MT achieved its optimal activity in presence of 1.0, 1.25, 2.0, 2.2 and 3.2 mM of pyruvic acid, acrylic acid, propanoic acid, 2-oxo-butanoic acid, and 2-oxo-octanoic acid, respectively. Contrarily, the cresolase activity of MT was inhibited by all type of the above acids. Propanoic acid caused an uncompetitive mode of inhibition (Ki=0.14 mM), however, the pyruvic, acrylic, 2-oxo-butanoic and 2-oxo-octanoic acids showed a competitive manner of inhibition with the inhibition constants (Ki) of 0.36, 0.6, 3.6 and 4.5 mM, respectively. So, it seems that, there is a physical difference in the docking of mono- and o-diphenols to the tyrosinase active site. This difference could be an essential determinant for the course of the catalytic cycle. Monophenols are proposed to bind only the oxyform of the tyrosinase. It is likely that the binding of acids occurs through their carboxylate group with one copper ion of the binuclear site. Thus, they could completely block the cresolase reaction, by preventing monophenol binding to the enzyme. From an allosteric point of view, n-alkyl acids may be involved in activation of MT catecholase reactions.

Inhibitory effects of phloridzin dihydrate on the activity of mushroom ( Agaricus bisporus) tyrosinase

Inhibitory effects of the water extracts of Lavendula sp. on mushroom tyrosinase activity

Activity, structural and stability changes of mushroom tyrosinase by sodium dodecyl sulfate

A novel monofunctional benzyldithiocarbamate, C(6)H(5)CH(2)NHCSSNa (I), and a bifunctional p-xylidine-bis(dithiocarbamate), NaSSCNHCH(2)C(6)H(4)CH(2)NHCSSNa (II), as sodium salts, were synthesized by reaction between p-xylylenediamine or benzylamine with CS(2) in the presence of NaOH. They were characterized by spectroscopic techniques such as (1)H NMR, IR, and elemental analysis. These water-soluble compounds were examined for their inhibition of both activities of mushroom tyrosinase (MT) from a commercial source of Agricus bisporus. l-3,4- Dihydroxyphenylalanine (L-DOPA) and l-tyrosine were used as natural substrates for the catecholase and cresolase enzyme reactions, respectively. Kinetic studies showed noncompetitive inhibition of I and mixed type inhibition of II on both activities of MT. The inhibition constant (K(I)) of II was smaller than that of I. Raising the temperature from 27 to 37 degrees C caused a decrease in K(I) values of I and an increase in values of II. The binding process for inhibition of I was only entropy driven, which means that the predominant interaction in the active site of the enzyme is hydrophobic; meanwhile, the electrostatic interaction can be important for the inhibition of II due to the enthalpy driven binding process. Fluorescence studies showed a decrease of emission intensity without a shift of emission maximum in the presence of different concentrations of compounds. An extrinsic fluorescence study did not show any considerable change of the tertiary structure of MT. Probably, the conformation of inhibitor-bound MT is stable and inflexible compared with uninhibited MT.

A novel monofunctional benzyldithiocarbamate, C6H5CH2NHCSSNa (I), and a bifunctional p-xylidine-bis(dithiocarbamate), NaSSCNHCH2C6H4CH2NHCSSNa (II), as sodium salts, were synthesized by reaction between p-xylylenediamine or benzylamine with CS2 in the presence of NaOH. They were characterized by spectroscopic techniques such as 1H NMR, IR, and elemental analysis. These water-soluble compounds were examined for their inhibition of both activities of mushroom tyrosinase (MT) from a commercial source of Agricus bisporus. l-3,4- Dihydroxyphenylalanine (L-DOPA) and l-tyrosine were used as natural substrates for the catecholase and cresolase enzyme reactions, respectively. Kinetic studies showed noncompetitive inhibition of I and mixed type inhibition of II on both activities of MT. The inhibition constant (KI) of II was smaller than that of I. Raising the temperature from 27 to 37°C caused a decrease in KI values of I and an increase in values of II. The binding process for inhibition of I was only entropy driven, which means that the predominant interaction in the active site of the enzyme is hydrophobic; meanwhile, the electrostatic interaction can be important for the inhibition of II due to the enthalpy driven binding process. Fluorescence studies showed a decrease of emission intensity without a shift of emission maximum in the presence of different concentrations of compounds. An extrinsic fluorescence study did not show any considerable change of the tertiary structure of MT. Probably, the conformation of inhibitor-bound MT is stable and inflexible compared with uninhibited MT.

Activity and structural changes of mushroom tyrosinase induced by n-alkyl sulfates

In this work, the inhibitory ability and mechanism of ω3‐nicotinic acid (ω3‐NA) and ω3‐Picolinic acid (ω3‐PA) complexes on the activity of mushroom tyrosinase (MT) were scrutinized for progressing a novel MT inhibitor. The complexes were synthesized. It was shown to have a considerable inhibition on the MT activity and K i value of ω3‐NA and ω3‐PA on MT equal to 5.2 and 5.1 mM, respectively. ω3‐NA and ω3‐PA inhibited MT with V max values in the range of 0.134 mM and 0.14 mM, respectively. The outputs obtained from fluorescence quenching specified that ω3‐NA and ω3‐PA could interact with MT. Especially, the decrease in fluorescence intensity was due to the formation of a ligand‐enzyme complex which was mostly motivated by hydrogen bonding and hydrophobic forces. The presence of ω3‐NA and ω3‐PA altered the structure of MT and reduced the α‐helix of the enzyme. Molecular docking investigation along with molecular dynamics simulation exhibited that ligands‐MT formation is directed by hydrogen binding with Trp136, His263, and Val299 residues. The results highlight that ω3‐NA and ω3‐PA can be considered as possible inhibitors in treating hyperpigmentation via MT enzyme inhibition.

Two structurally related compounds, phenyl dithiocarbamate sodium salt (I) and p-phenylene-bis (dithiocarbamate) sodium salt (II) were prepared by reaction of the parent aniline and p-phenylenediamine with CS₂ in the presence of sodium hydroxide. These water soluble compounds were characterized by spectroscopic techniques, IR, ¹H NMR and elemental analysis. The inhibitory effects of both compounds on both activities of mushroom tyrosinase (MT) from Agricus bisporus were studied at two temperatures, 27°C and 37°C. L-3, 4-dihydroxyphenylalanine (L-DOPA), and l-tyrosine were used as natural substrates for the catecholase and cresolase enzyme reactions, respectively. Kinetic analysis confirmed noncompetitive inhibition mode of I and mixed type of II on both activities of MT; I and II inhibit MT with inhibition constants (K(I)) of 300 µM and 4 µM, respectively. Analysis of thermodynamic parameters indicated predominant involvement of hydrophobic interactions in binding of I and electrostatic ones in binding of II to MT. It seems that II is a more potent MT inhibitor due to its two charged head groups able to chelate copper ions in the enzyme active site. Intrinsic fluorescence studies as a function of concentrations of both compounds showed unexpectedly quenching of emission intensity without any shift of emission maximum. Extrinsic ANS-fluorescence indicated that only binding of I induces limited changes in the tertiary structure of MT, in agreement with the postulated hydrophobic nature of the binding mechanism.

Modifying effects of carboxyl group on the interaction of recombinant S100A8/A9 complex with tyrosinase

Mushroom tyrosinase (MT) structural changes in the presence of <TEX>$Cu ^{2+}$</TEX> and <TEX>$Ni ^{2+}$</TEX> were studied separately. Far-UV CD spectra of the incubated MT with the either of the metal ions indicated reduction of the well-ordered secondary structure of the enzyme. Increasing in the maximum fluorescence emission of anilinonaphthalene-8-sulfonic acid (ANS) was also revealing partial unfolding caused by the conformational changes in the tertiary structure of MT. Thermodynamic studies on the chemical denaturation of MT by dodecyl trimethylammonium bromide (DTAB) showed decrease in the stability of MT in the presence of <TEX>$Cu ^{2+}$</TEX> or <TEX>$Ni ^{2+}$</TEX> using their activation concentrations. Both activities of MT were also assessed in the presence of different concentrations of these ions, separately, with various monophenols and their corresponding diphenols. Kinetic studies revealed that cresolase activity on p-coumaric acid was boosted in the presence of either of the metal ions, but inhibited when phenol, L-tyrosine, or 4-[(4-methylphenyl)azo]-phenol was substrate. Similarly, catecholase activity on caffeic acid was enhanced in the presence of <TEX>$Cu ^{2+}$</TEX> or <TEX>$Ni ^{2+}$</TEX>, but inhibited when catechol, L-DOPA, or 4-[(4-methylbenzo)azo]-1,2-benzenediol was substrate. Results of this study suggest that both cations make MT more fragile and less active. However, the effect of the substrate structure on the MT allosteric behavior can not be ignored.

Successful resonance Raman study of cresolase activity of mushroom tyrosinase