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An extracellular laccase (GLL) was purified from fermentation broth of the litter-decomposing fungus Gymnopus luxurians by four chromatography steps, which resulted in a high specific activity of 118.82 U/mg, purification fold of 41.22, and recovery rate of 42.05%. It is a monomeric protein with a molecular weight of 64 kDa and N-terminal amino acid sequence of AIGPV TDLHI, suggesting that GLL is a typical fungal laccase. GLL demonstrated an optimum temperature range of 55°C–65°C and an optimum pH 2.2 toward 2,2′-azino-bis(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS). It displayed considerably high thermostability and pH stability with about 63% activity retained after 24 h at 50°C, and 86% activity retained after 24 h at pH 2.2, respectively. GLL was significantly enhanced in the presence of K+, Na+, and Mg2+ ions. It demonstrated Km of 539 μM and kcat/Km of 140 mM–1⋅s–1 toward ABTS at pH 2.2 and 37°C. Acetosyringone (AS) and syringaldehyde (SA) were the optimal mediators of GLL (0.4 U/ml) for dye decolorization with decolorization rates of about 60%–90% toward 11 of the 14 synthetic dyes. The optimum reaction conditions were determined to be mediator concentration of 0.1 mM, temperature range of 25°C –60°C, and pH 4.0. The purified laccase was the first laccase isolated from genus Gymnopus with high thermostability, pH stability, and effective decolorization toward dyes, suggesting that it has potentials for textile and environmental applications.

The present study aims to investigate the effects of ginsenoside Rg3 combined with oxaliplatin on the proliferation and apoptosis of hepatocellular carcinoma cells and the related mechanism. In this study, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was applied to examine the proliferation rate of hepatocellular carcinoma cell SMMC-7721 with different treatment. Flow cytometry was performed to examine apoptosis rate of hepatocellular carcinoma cells with different treatment. Immunofluorescence and Western blot methods were used to evaluate the expressions of proliferating cell nuclear antigen (PCNA) and cyclin D1 in different groups. We found that ginsenoside Rg3, oxaliplatin or ginsenoside Rg3 + oxaliplatin significantly suppressed the proliferation and promoted the apoptosis of SMMC-7721. Meanwhile, ginsenoside Rg3, oxaliplatin or ginsenoside Rg3 + oxaliplatin also significantly inhibited the expressions of PCNA and cyclin D1. Moreover, compared with ginsenoside Rg3 group and oxaliplatin group, the effect of ginsenoside Rg3 + oxaliplatin was more remarkable. Taken together, cells treated with oxaliplatin+ ginsenoside enhanced the anti-tumor effect and may inhibit the proliferation and promoted apoptosis of hepatocellular carcinoma via regulating the expression of PCNA and cyclin D1.

A novel laccase was purified from fermentation broth of white rot fungus Trametes sp. LAC-01 using an isolation procedure involving three ion-exchange chromatography steps on DEAE-cellulose, SP-Sepharose, and Q-Sepharose, and one gel-filtration step. The purified enzyme (TSL) was proved as a monomeric protein with a Mr of 59kDa based on SDS-PAGE and FPLC. Partial amino acid sequences were obtained by LC-MS/MS sharing considerably high sequence similarity with that of other laccases. It possessed optimal pH of 2.6 and temperature of 60°C using ABTS as the substrate. The Km of the laccase toward ABTS was estimated to 30.28μM at pH 2.6 and 40°C. TSL manifested considerably high oxidizing activity toward ABTS, but was avoid of degradative activity toward benzidine, caftaric acid, etc. It was effective in the decolorization of phenolic dyes - Bromothymol Blue and Malachite Green with decolorization rate higher than 60% after 24h of incubation. Adjunction of Cu(2+) with the final concentration of 2.0mmol/L significantly activated laccase production with a steady high level of 275.8-282.2U/mL in 96-144h. The high yield and short production period makes Trametes sp. LAC-01 and TSL potentially useful for industrial and environmental application and commercialization.

NAC transcription factor is a new-found and multifunctional transcription factor in plants. The purpose of the paper is to study the relative expression trends of BnNAC transcription factor gene under cold (4°C), salinity (200 mM NaCl) and abscisic acid (ABA) (100 μM) stress in Brassica napus. The functional region of BnNAC gene was cloned through the homologous cloning method. The sequences of nucleic acid and amino acid were analyzed using BLAST and DNAMAN software. Relative expression trends of BnNAC gene were detected by applying real-time relative quantification PCR (RT-qPCR) under cold, salinity and ABA treatment. A 440 bp BnNAC gene segment was acquired in Brassica napus. BnNAC gene expressions of Brassica napus seedlings were induced under cold, salinity and ABA treatment conditions. The results of RT-qPCR analysis revealed that, under salinity, cold and ABA conditions, the relative expression of BnNAC gene increases with time, reaches the peak at 24h, 3h, 12h respectively and then decreases. Our research showed that BnNAC plays a positive role in salinity, cold and ABA stress response.

The development of the stone and formation of peach (Prunus persica) fruit were explored in this work using a proteomic approach. Sixty-eight proteins with different expression patterns were identified in both the endocarp and mesocarp during early fruit development (from 28 to 59 days after flowering) and the majority were involved in primary or secondary metabolism. In contrast to most proteins associated with primary metabolism in the endocarp, whose expression is down-regulated, expression of pyruvate dehydrogenase (PDH) unexpectedly increased exponentially. Moreover, its expression pattern was linearly positively correlated with the exponentially growing lignin content (R = 0.940), which suggests that PDH may play a role in endocarp lignification. Our data also revealed different spatiotemporal expressions of enzymes involved in the lignin and flavonoid pathways that provided proteome-level evidence to support the hypothesis that these two pathways are competitive during endocarp development. In addition, we observed endocarp-specific oxidative stress and propose that it may act as a stimulating factor in activating lignification and subsequent programmed cell death in the endocarp.

Deterioration of peach fruit quality because of split-pit formation had bad influence on market (1). Though many studies have mentioned the mechanism of split-pit formation, further studied are still needed to uncover the development mechanism of endocarp cell during pit hardening process, especially the Programmed cell death (PCD) process of peach endocarp cell. Here, the development of the peach endocarp cell from the two Chinese peach varieties "Jingyu" and "Okubo" was detailedly studied to understand the PCD of endocarp cell during the lignin formation process. According to our morphology observation using transmission electron microscopy, the PCD process of the endocarp cell, characterized by vacuolar enlargement, might have been initiated as early as eight days after full athesis (maybe even more earlier), which is earlier than that of the split-pit formation process. In our studies, almost all the endocarp cell organelles were found to degenerate at forty days after full athesis, when thickening of the endocarp cell wall was also observed. Furthermore, the involvement of hydrolytic enzymes, in particular proteases and nucleases, in the autolytic process of PCD of peach endocarp cells was quantitatively analyzed during pit hardening process. The peach endocarp protein was found to be rapidly degraded by newly expressed proteases since 35 days after full athesis. Elevation of nuclease activities was found to be also coupled with PCD of peach endocarp cells. Furthermore, DNA degradation was found to happen earlier than RNA degradation and hydrolyzing of endocarp soluble protein during the peach pit hardening process.

Few studies were mentioned to illustrate the role of plant hormones-IAA and ethylene in split-pit formation. Here, a random sample of 100 developing fruits from 7-year-old early maturing nectarines (Prunus persica L. Batsch cv. Ruiguang 5) was used as test material. The shape index and growth index of the sampled nectarines was comparatively analyzed to show the relation between fruit un-uniform growth and their split-pit formation. The split-pit fruits were found to have a higher degree of un-uniform growth than that of the normal fruits, which were demonstrated by higher ratio of transverse diameter/lateral diameter and bigger difference between the flesh thickness of the two sides in the middle part of the fruits (along the line of the transverse diameter) about twelve weeks after full anthesis (p < 0.05). Subsequently, a higher degree of unbalanced distribution of IAA and especially ethylene was observed in the split-pit fruits compared to the normal fruits, which may correlate with un-balanced expansion of the nectarines flesh. Higher concentration of IAA and especially ethylene was found in the middle part of the split-pit fruits than those in the sutures, while those in the normal fruits were not significant. Furthermore, the caliosity was specifically observed in the internal surface of the split-pit nectarines endo carp as early as four weeks after full anthesis, but not in the normal nectarines. The occurrence of caliosit in the endo carp may be an early signal of spilt-pit.