Purification and Characterization of a Naringinase from Cryptococcus albidus

Abstract

Naringinase which was extracted from the fermented broth of Cryptococcus albidus was purified about 42-folds with yield 0.7% by sulfate fractionation and chromatography on Toyopearl HW-60, Fractogel DEAE-650-s, and Sepharose 6B columns. Molecular weight of protein determined by gel filtration and SDS-PAGE was 50 kDa. Naringinase of C. albidus includes high content of the dicarbonic and hydrophobic amino acids. Enzyme contains also carbohydrate component, represented by mannose, galactose, rhamnose, ribose, arabinose, xylose, and glucose. The enzyme was optimally active at pH 5.0 and 60 °C. Naringinase was found to exhibit specificity towards p-nitrophenyl-α-L-rhamnose, p-nitrophenyl-β-D-glucose, naringin, and neohesperidin. Its K towards naringin was 0.77 mM and the V was 36 U/mg. Naringinase was inhibited by high concentrations of reaction product-L-rhamnose. Enzyme revealed stability to 20% ethanol and 500 mM glucose in the reaction mixture that makes it possible to forecast its practical use in the food industry in the production of juices and wines.

The influence of coordinative tartrate and malatogermanate compounds on the activity of ?-L-rhamnosidase preparations from Penicillium tardum, Eupenicillium erubescens and Cryptococcus albidus

“…, (7) was also observed at concentration 0.01%. A decrease in the inhibitory effect with an increase in the exposure time was also observed, similar to α-Lrhamnosidase from P. tardum.…”

“…Compounds (7) and (9) at both concentrations and exposure time 0.5 h did not affect the activity of the studied α-L-rhamnosidase (activity was at the control level). All other compounds inhibited the activity of α-L-rhamnosidase from P. tardum.…”

“…Previously, based on inhibitory and kinetic analysis with Dixon and Lineweaver-Burk plots and using group-specific reagents, we revealed some active functional groups in α-L-rhamnosidases from P. tardum, e. erubescens and C. albidus [7][8][9]. A significant inhibition (more than 50%) of the activity of all the studied enzymes by 1-[3-(dimethylamino) propyl]-3-ethylcarbodiimide methiodide may indicate an important role of carboxyl groups of aspartic or glutamine amino acids in the catalytic activity of the enzymes.…”

“…All compounds were comprehensively characterized by modern physical and chemical methods: elemental analysis, infrared spectroscopy, thermogravimetric analysis, single crystal XRD. Structure data of the synthesized compounds were presented in publications [20][21][22] and deposited in the Cambridge Crystallographic Database: 1878103 (2), 1883675 (3), 1883676 (4), 1513407 (5), 1513408 (6), 1576554 (7). Representations of the studied compounds are shown in Fig.…”

“…We selected the most active strains -Penicillium tardum [4], eupenicillium erubescens [5] and Cryptococcus albidus [6] and obtained α-L-rhamnosidase preparations from their culture fluids. Then we studied their physiochemical properties, substrate specificity, and functional groups involved in the catalytic process [7][8][9]. To obtain highly productive microbial enzymes, e.g.…”