Biomedical subjects

C Lindbladh

Publications and source records attributed to C Lindbladh.

25 records · Page 2Linked to original sources

PubMed1984

Inactivation of human tyrosinase by cysteine. Protection by dopa and tyrosine.

Human tyrosinase prepared from cultured melanoma cells is inactivated by 10 mM cysteine. The inactivation of the enzyme by cysteine is less pronounced in the presence of catalase and superoxide dismutase. Thus, oxygen radicals and/or hydrogen peroxide may contribute to the inactivation of human tyrosinase by cysteine. Dopa and/or tyrosine protects tyrosinase against inactivation by cysteine. The protection observed with tyrosine alone indicates that oxidation of substrate is not necessary for the protection. The effect of dopa and/or tyrosine is probably due to steric hindrance at the active site preventing the access of cysteine to the copper.

Catalase↗

PubMed1983

Tyrosinase activity in the medium of human melanoma cell cultures.

The medium of cultured melanoma cells was studied for tyrosine hydroxylation and dopa-oxidizing activity. The supernatant obtained after centrifugation at 100 000 g for 2 hours was treated with ammonium sulphate, and the precipitate obtained between 35 and 50% saturation was used. Dopa was determined as the product of tyrosine hydroxylation and 5-S-cysteinyldopa as the product of dopa oxidase activity. Determinations were performed with HPLC and electrochemical detection. Our preparation of culture medium of cells showed the following. 1) No hydroxylation of tyrosine in the absence of co-factor. 2) Hydroxylation of L-tyrosine in the presence of dopamine. No hydroxylation with boiled medium. Minimal effect of catalase on hydroxylation. 3) Hydroxylation of tyrosine in the presence of ascorbic acid. Hydroxylation was catalyzed also with boiled medium. Catalase strikingly diminished hydroxylation. 4) Oxidation of L-dopa to dopaquinone determined as its main reaction product with cysteine, 5-S-cysteinyl-dopa. There was negligible oxidation with boiled medium. 5) With dopamine as co-factor the catalysis of tyrosine hydroxylation was stereospecific for L-tyrosine. Dopa oxidase activity was also stereospecific for L-dopa.

Ascorbic Acid↗

PubMed1983

The effect of catalase on the inactivation of tyrosinase by ascorbic acid and by cysteine or glutathione.

When tyrosine was incubated with tyrosinase in the presence of ascorbic acid, dopa and 5-hydroxydopa were formed and the enzyme was inactivated. In the presence of catalase, more dopa and 5-hydroxydopa were formed because enzyme inactivation was prevented. Incubation of dopa and cysteine with small amounts of mushroom tyrosinase led to rapid inactivation of the enzyme. This inactivation was accelerated in the presence of catalase. New systems developed have been useful in demonstrating the role of hydrogen peroxide in tyrosinase inactivation by several compounds of importance in melanin biochemistry. Cysteine and glutathione inactivated tyrosinase. Addition of catalase increased the inactivation at high thiol concentrations, but decreased the inactivation at low concentrations. Ascorbic acid and 5-hydroxydopamine also inactivated tyrosinase, but with these compounds inactivation was completely prevented by addition of catalase. The inactivation by dopamine was negligible under the experimental conditions. Inactivation of tyrosinase by ascorbic acid and by 5-OH-dopamine was found dependent on oxygen, whereas inactivation by cysteine and glutathione was independent of oxygen. Large amounts of serum albumin protected tyrosinase from inactivation by ascorbic acid and 5-OH-dopamine, but did not prevent inactivation by cysteine and glutathione. The presence of substrate had a protective effect on the inactivation of tyrosinase by cysteine.

Ascorbic Acid↗

PubMed1983

Dopa oxidation and tyrosine oxygenation by human melanoma tyrosinase.

A tyrosinase purified from cultured human melanoma cells was studied for dopa oxidation and tyrosine oxygenation. Km for oxidation of L-dopa was 0.5 mM, and for D-dopa 3 mM. L-tyrosine was oxygenated only in the presence of a cosubstrate. L-Dopa was superior to D-dopa, dopamine, L- and D-alpha-methyldopa, dopac, and 5,6-dihydroxyindole-2-carboxylic acid as cosubstrate. Ascorbic acid, 5-S-cysteinyldopa, and 5-OH-dopa did not function as cosubstrates. The rate of tyrosine hydroxylation was much lower than that of dopa oxidation. Tyrosine inhibits dopa oxidation, and dopa in high concentrations inhibits tyrosine hydroxylation. The cosubstrate function of dopa, the substrate functions of dopa and tyrosine, and the mutual inhibition of tyrosinase by these compounds are explained in three equations.

Catechol Oxidase↗

PubMed1978

Trichochromes in the urine of melanoma patients.

The urine of patients with melanoma metastases and increased urinary excretion of 5-S-cysteinyldopa was examined for trichochromes. Five of 16 patients showed urinary excretion of trichochromes B and C. None excreted trichochromes E or F. All patients showing trichochrome excretion had very large amounts of cysteinyldopa in their urine.

Adult↗

PubMedDate not supplied

Selection of a cyclic nonapeptide inhibitor to alpha-chymotrypsin using a phage display peptide library.

A cyclic nonapeptide library displayed on filamentous bacteriophages was selected 6 times against alpha-chymotrypsin (EC 3.4.21.1) at three different pH conditions (6.5, 7.0, and 7.5). Phage peptide clones from the sixth selection, at all three pH conditions, interacted more strongly with alpha-chymotrypsin than the original library and a wild-type phage did. DNA sequencing of the selected phage peptide clones showed that different cyclic nonapeptide sequences had been selected at the different pH conditions. The oxidized form of the synthetic peptide, Cys-Cys-Phe-Ser-Trp-Arg-Cys-Arg-Cys, selected at pH 7.5, could completely inhibit the enzymatic activity of alpha-chymotrypsin. The structurally related enzymes trypsin (bovine) and elastase (porcine) were only marginally inhibited by the same peptide under the same conditions. The inhibition constant for alpha-chymotrypsin was estimated to be 10(-6) M. Phage clones expressing this peptide had a lower affinity for phenylmethylsulfonylfluoride-modified alpha-chymotrypsin than for natural alpha-chymotrypsin as determined by an enzyme immunosorbent assay. This peptide phage clone was also competitively prevented from binding to alpha-chymotrypsin by the corresponding synthetic oxidized peptide. Collectively, the results suggest that the oxidized form of the selected peptide Cys-Cys-Phe- Ser-Trp-Arg-Cys-Arg-Cys interacts with the active site of alpha-chymotrypsin and acts as a specific inhibitor to the enzyme. To our knowledge, the selected sequence Cys-Cys-Phe-Ser-Trp-Arg-Cys-Arg-Cys has not been found in nature.

Amino Acid Sequence↗

PubMedDate not supplied

Dose-response protective effect of salbutamol on methacholine airway responsiveness using pressurized metered dose inhalers and Turbuhalers.

The purpose of this study was to estimate the relative dose potency of salbutamol Turbuhaler compared with salbutamol pressurized metered dose inhaler (pMDI) with respect to the protective effect against methacholine bronchoconstriction. Twenty-three asthmatic subjects with stable asthma participated in the study. Baseline forced expiratory volume in 1 s (FEV1) was 70% or more of predicted, and baseline methacholine provocative concentration causing a 20% fall in FEV1 (PC20) was 4 mg/mL or less. The design was randomized, double-blind, double-dummy, crossover and placebo controlled and was conducted over seven study days. On each study day, the subjects inhaled 50 microg or 100 microg of salbutamol via Turbuhaler, 100 microg, 200 microg, 400 microg or 800 microg of salbutamol via pMDI, or placebo in randomized order. PC20 was determined 30 mins after inhalation. Increasing doses of salbutamol pMDI increased the PC20 in a dose-dependent fashion from 3.9 mg/mL after placebo to 13.3 mg/mL after pMDI 100 microg, 19.0 mg/mL after 200 microg, 32.6 mg/mL after 400 microg, and 35.1 mg/mL after 800 microg. The half-maximum response dose for pMDI (ED50) was 104 microg. Salbutamol Turbuhaler 50 microg increased the PC20 to 10.0 mg/mL and 100 microg to 12.6 mg/mL. Salbutamol pMDI 200 microg provided significantly greater protection to methacholine than pMDI 100 microg or Turbuhaler 100 microg and significantly less protection than pMDI 400 microg (P<0. 05). This study demonstrates that the relative protective dose potency of inhaled beta-agonists can be determined by comparing their effects on methacholine airway responsiveness. The estimated relative protective dose potency for salbutamol Turbuhaler in comparison with pMDI was 1.38 (95% CI 0.67 to 2.87) at 50 microg and was 0.96 (95% CI 0.56 to 1.64) at 100 microg.

Adrenergic beta-Agonists↗