Purification of a novel low-molecular-mass laccase with HIV-1 reverse transcriptase inhibitory activity from the mushroom Tricholoma giganteum.
A laccase with a novel N-terminal sequence, a low molecular mass of 43 kDa smaller than those of previously reported laccases, a pH optimum of 4, and a temperature optimum at 70 degrees C was isolated from fresh fruiting bodies of the mushroom Tricholoma giganteum. The activity of the enzyme rose steadily from 20 to 50 degrees C, increased very slowly from 50 to 70 degrees C, and fell slightly when the temperature was further increased to 80 degrees C. The activity of the laccase underwent little changes over the pH range 3.0-5.0. However, the enzyme activity dwindled to nothing after exposure to 100 degrees C for 10 min and when the ambient pH was 7 or above. The procedure used for purifying the enzyme included ion exchange chromatography on DEAE-cellulose, affinity chromatography on Affi-gel blue gel, ion exchange chromatography on CM-cellulose, and FPLC-gel filtration on Superdex 75. The laccase was unadsorbed on DEAE-cellulose and adsorbed on Affi-gel blue gel and CM-cellulose. It inhibited HIV-1 reverse transcriptase with an IC(50) of 2.2 microM.
Wang
H X
HX
Department of Microbiology, China Agricultural University, and State Key Laboratory of Agrobiotechnology, Beijing, China.
Ng
T B
TB
D000363:Agaricales / Q000378:metabolism*; D002482:Cellulose / Q000737:chemistry; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D015497:HIV-1 / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D020128:Inhibitory Concentration 50; D042845:Laccase / Q000737:chemistry / Q000302:isolation & purification*; D010936:Plant Extracts; D017434:Protein Structure, Tertiary; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology*; D013696:Temperature
D010936:Plant Extracts; D018894:Reverse Transcriptase Inhibitors; D002482:Cellulose; D042845:Laccase