Heterologous extracellular expression and initial characterization of the peroxisomal catalase from the methylotrophic yeast Hansenula polymorpha in Pichia pastoris

Catalase is well known to eliminate H₂O₂ in cells and reduces the toxicity of peroxide compounds. A catalase gene HpCAT1 of methylotrophic yeast Hansenula polymorpha without the part coding the native signal peptide was cloned into expression vector pYM3165 and then integrated into genome of Pichia pastoris GS115 by electroporation. The result of the enzyme activity assay and SDS-PAGE demonstrated that the recombinant protein (HpCAT1) of H. polymorpha was extracellularly expressed in P. pastoris. The expressed catalase was recovered from the culture supernatant of P. pastoris GS115 and purified by (NH₄)₂SO₄ fractionation and Ni-NTA affinity chromatography. The main biochemical properties of the recombinant protein HpCAT1, such as thermodependence and thermostability, pH optimum and pH stability, as well as the effect of metal ions and chemicals, were characterized. With H₂O₂ as the substrate, HpCAT1 displayed pH and temperature optima of ～2.6 and 45°C, respectively. The recombinant HpCAT1 activity was inhibited by 1 mM Hg²⁺ and Cu²⁺, but was highly enhanced by 1.0 mM Fe²⁺.     

A novel melatonin agonist Neu-P11 facilitates memory performance and improves cognitive impairment in a rat model of Alzheimer' disease

Previous studies have shown that melatonin is implicated in modulating learning and memory processing. Melatonin also exerts neuroprotective activities against Aβ-induced injury in vitro and in vivo. Neu-P11 (piromelatine, N-(2-(5-methoxy-1H-indol-3-yl)ethyl)-4-oxo-4H-pyran-2-carboxamide) is a novel melatonin (MT1/MT2) receptor agonist and a serotonin 5-HT1A/1D receptor agonist recently developed for the treatment of insomnia. In the present study we firstly investigated whether Neu-P11 and melatonin enhance memory performance in the novel object recognition (NOR) task in rats, and then assessed whether Neu-P11 and melatonin improve neuronal and cognitive impairment in a rat model of Alzheimer' disease (AD) induced by intrahippocampal Aβ_(1–42) injection. The results showed that a single morning or afternoon administration of Neu-P11 enhanced object recognition memory measured at 4 or 24 h after training. Melatonin was effective in the memory facilitating effects only when administered in the afternoon. Further results showed that intrahippocampal Aβ_(1–42) injection resulted in hippocampal cellular loss, as well as decreased learning ability and memory in the Y maze and NOR tasks in rats. Neu-P11 but not melatonin attenuated cellular loss and cognitive impairment in the rat AD model. The current data suggest that Neu-P11 may serve as a novel agent for the treatment of AD.     

The Arabidopsis oligopeptidases TOP1 and TOP2 are salicylic acid targets that modulate SA‐mediated signaling and the immune response

Salicylic acid (SA) is a small phenolic molecule with hormonal properties, and is an essential component of the immune response. SA exerts its functions by interacting with protein targets; however, the specific cellular components modulated by SA and critical for immune signal transduction are largely unknown. To uncover cellular activities targeted by SA, we probed Arabidopsis protein microarrays with a functional analog of SA. We demonstrate that thimet oligopeptidases (TOPs) constitute a class of SA‐binding enzymes. Biochemical evidence demonstrated that SA interacts with TOPs and inhibits their peptidase activities to various degrees both in vitro and in plant extracts. Functional characterization of mutants with altered TOP expression indicated that TOP1 and TOP2 mediate SA‐dependent signaling and are necessary for the immune response to avirulent pathogens. Our results support a model whereby TOP1 and TOP2 act in separate pathways to modulate SA‐mediated cellular processes.     

Synthesis and antitumor activities of novel α-aminophosphonates dehydroabietic acid derivatives

A series of novel α-aminophosphonate derivatives containing DHA structure were designed and synthesized as antitumor agents. In vitro antitumor activities of these compounds against the NCI-H460 (human lung cancer cell), A549 (human lung adenocarcinoma cell), HepG2 (human liver cancer cell) and SKOV3 (human ovarian cancer cell) human cancer cell lines were evaluated and compared with commercial anticancer drug 5-fluorouracil (5-FU), employing standard MTT assay. The pharmacological screening results revealed that many compounds exhibited moderate to high levels of antitumor activities against the tested cancer cell lines and that most demonstrated more potent inhibitory activities compared with the commercial anticancer drug 5-FU. The action mechanism of representative compound 7c was preliminarily investigated by acridine orange/ethidium bromide staining, Hoechst 33258 staining, JC-1 mitochondrial membrane potential staining and flow cytometry, which indicated that the compound can induce cell apoptosis in NCI-H460 cells. Cell cycle analysis showed that compound 7c mainly arrested NCI-H460 cells in G1 stage.     

Synthesis and acrosin inhibitory activities of 5-phenyl-1H-pyrazole-3-carboxylic acid amide derivatives

A series of novel 5-phenyl-1H-pyrazole-3-carboxylic acid amide derivatives were designed, synthesized, and their acrosin inhibitory activities in vitro were evaluated. The results of the acrosin inhibitory activity showed that all target compounds were more potent than control TLCK. Compounds AQ-A1, AQ-D3, AQ-D4, AQ-E4 and AQ-E5 exhibited stronger acrosin inhibitory activities than control ISO-1. Especially, compound AQ-E5 displayed the most potent acrosin inhibitory activity in all the compounds, with an IC₅₀ of 0.01 μmol/mL. This study provided a new structural class for the development of novel acrosin inhibitory agents.     

Conformation constraint of anilides enabling the discovery of tricyclic lactams as potent MK2 non-ATP competitive inhibitors

Conformation restriction of linear N-alkylanilide MK2 inhibitors to their E-conformer was developed. This strategy enabled rapid advance in identifying a series of potent non-ATP competitive inhibitors that exhibited cell based activity in anti-TNFα assay.     

马哈利樱桃大孢子发生和雌配子体的发育

采用石蜡切片法对马哈利樱桃大孢子发生和雌配子体发育过程进行观察研究。结果表明:(1)马哈利樱桃雌配子体发育早期,在单室子房内可以看到2个倒生胚珠,但在后期其中一个退化,另一个发育为种子;其胚珠具双珠被,为厚珠心。(2)大孢子母细胞减数分裂形成直线型四分体,功能大孢子位于合点端;胚囊发育为蓼型,成熟胚囊为七细胞八核。(3)根据不同时间花的外部形态特征与内部解剖学对比的观察结果,在陕西关中地区,三月下旬是马哈利樱桃雌性生殖细胞分化和发育的重要时期,果园在此期间应加强肥水管理。     

7个大穗型小麦-黑麦代换系间杂交后代的形态学与SSR分析

对从小麦-黑麦代换系5R/5A与1R/1D杂交后代中选育的大穗型品系1-5、1-6、1-7、1-8、1-9、1-10、1-11进行形态学与SSR分析。结果表明,7个品系田间表现遗传性稳定,且具有黑麦大穗、抗病等优良性状;利用黑麦特异引物PSC119.1确定7个品系均导入了黑麦染色体片段,引物SCM138扩增结果表明,在1-6、1-7、1-8、1-9、1-10中导入了黑麦染色体5RS片段,引物SCM120、TSM604可以在7个品系中稳定扩增出黑麦5R长臂和1R短臂特异片段。以上结果可为小麦遗传育种与品质改良提供基础材料。