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. 2012:2012:536725.
doi: 10.1155/2012/536725. Epub 2012 Mar 21.

A laccase with HIV-1 reverse transcriptase inhibitory activity from the broth of mycelial culture of the mushroom Lentinus tigrinus

LiJing Xu  1 , HeXiang Wang, TziBun Ng
Affiliations

A laccase with HIV-1 reverse transcriptase inhibitory activity from the broth of mycelial culture of the mushroom Lentinus tigrinus

LiJing Xu et al. J Biomed Biotechnol. 2012.

Abstract

A 59 kDa laccase with inhibitory activity against HIV-1 reverse transcriptase (IC(50) = 2.4 μM) was isolated from the broth of mycelial culture of the mushroom Lentinus tigrinus. The isolation procedure involved ion exchange chromatography on DEAE-cellulose and CM-cellulose, and gel filtration by fast protein liquid chromatography on Superdex 75. The laccase was adsorbed on both types of ion exchangers. About 95-fold purification was achieved with a 25.9% yield of the enzyme. The procedure resulted in a specific enzyme activity of 76.6 U/mg. Its N-terminal amino acid sequence was GIPDLHDLTV, which showed little similarity to other mushroom laccase and other Lentinus tigrinus strain laccase. Its characteristics were different from previously reported laccase of other Lentinus tigrinus strain. Maximal laccase activity was observed at a pH of 4 and at a temperature of 60°C, respectively. This study yielded the information about the potentially exploitable activities of Lentinus tigrinus laccase.

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Figures

Figure 1
Figure 1
Ion exchange chromatography on CM-cellulose column (2.5 × 20 cm). Sample: fraction of Lentinus tigrinus incubation broth that was adsorbed on DEAE-cellulose. Fraction size: 8 mL. Starting buffer: 10 mM NH4OAc buffer (pH 4.5). Dotted line across right-hand side of chromatogram indicates linear NaCl concentration (0-1 M) gradient used to CM2 and CM3.
Figure 2
Figure 2
FPLC gel filtration on Superdex 75 hR 10/30 column. Sample: fraction CM2. Buffer: 0.2 M NH4HCO3 buffer (pH 8.5). Fraction size: 0.8 mL. Flow rate: 0.4 mL/min.
Figure 3
Figure 3
Sodium dodecyl sulfate-polyacrylamide gel electrophoresis results. Left lane: molecular mass markers from Ge Healthcare. From top downward: phosphorylase b (94 kDa), bovine serum albumin (67 kDa), ovalbumin (43 kDa), carbonic anhydrase (30 kDa), soybean trypsin inhibitor (20 kDa), and α-lactalbumin (14.4 kDa). Right lane: purified laccase (fraction SU1).
Figure 4
Figure 4
Effect of pH on activity of Lentinus tigrinus laccase.
Figure 5
Figure 5
Effect of temperature on activity of Lentinus tigrinus laccase.
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