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国家自然科学基金(31370104)

作品数:6 被引量:16H指数:3
相关作者:张姜黄嘉丰武翠玲何海伦刘丹更多>>
相关机构:中南大学长治医学院中国科学院更多>>
发文基金:国家自然科学基金国家级星火计划更多>>
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Salinivibrio sp.YH4胞外蛋白酶EYHⅠ的分离纯化、酶学性质及基因克隆被引量:3
2016年
从中度嗜盐菌Salinivibrio sp.YH4发酵的粗酶液中分离纯化出蛋白酶EYHⅠ,对其进行酶学性质分析、串联质谱鉴定及全基因克隆。结果表明,EYHⅠ属于金属蛋白酶,最适温度55℃,热稳定性较好。最适p H为9.0,碱性条件下较稳定;在4 mol/L的Na Cl溶液中EYHⅠ仍保持较高活性,EYHⅠ全基因序列共1 836 bp,蛋白序列共611个氨基酸。比对发现EYHⅠ氨基酸序列与Salinivibrio sp.AF-2004所产Zn金属蛋白酶前体(ABI93183)同源性最高(96%)。结构分析表明,EYHⅠ由一个FTP结构域,一个Pep SY结构域,一个M4中性蛋白酶和一个PPC结构域组成。本研究为嗜盐菌及其胞外蛋白酶的生产和应用奠定了理论基础。
武翠玲刘丹杨兴昊吴日帮黄嘉丰张姜伦梓丰何海伦
关键词:蛋白酶分离纯化酶学性质基因克隆
The extraction of effective components and an antioxidant activity study of Tulipa edulis
2024年
Plant extracts from natural sources are an excellent choice for food additives and natural antioxidants.In this study,the active components of Tulipa edulis were extracted and analysed,and their antioxidant capacity was measured.Then,the crude extract mixture was separated and purified using a Sephadex LH-20 gel,and the antioxidant activity of the purified products was determined.Human umbilical vein endothelial human umbilical vein endothelial cells(HUVEC)cells were treated with 35 mmol/L glucose to construct a model of oxidative stress.Then,the cells were treated with the active component to observe whether the products of T.edulis could have a good protective effect on HUVEC cells induced by glucose.Transcriptome analysis was also performed on HUVEC cells after same treatment to explore the possible mechanism of the component F2 protecting HUVEC cells from oxidative stress induced by high glucose.The results showed that component F2 obtained from T.edulis has strong antioxidant activity.Moreover,F2 can play a strong antioxidant protective role in HUVEC cells.Meanwhile,the gene expression of heme oxygenase 1(HO-1),γ-glutamyl cysteine ligase catalytic subunit(GCLC)and NAD(P)H quinone oxidoreductase-1(NQO1)in HUVEC cells was up-regulated after treated with F2.This study provides reference value for the further development and application of T.edulis and the d evelopment of functional food.
Doudou ZhangDong XiaoTingting YinShuangzhi ZhaoOlena ZhurXun XiaoHailun HeLeilei Chen
关键词:PURIFICATIONANTIOXIDANT
西南地区高山湖泊中可培养细菌多样性及其所产胞外活性物质的特性被引量:4
2017年
【目的】研究西南不同地区的高山湖泊中可培养细菌的多样性及其产胞外蛋白酶、纤维素酶和胞外多糖的能力。【方法】以西南4个不同地区的高山湖泊:雷波的马湖(LB)、中缅边境的凯邦亚湖(ZM)、沙德的莲花湖(SD)、腾冲的青海湖(TC)的水样为研究对象,利用稀释涂布平板方法对可培养细菌进行分离筛选,然后通过对可培养细菌的生理生化指标和16S r RNA基因序列进行分析,初步确定细菌属别;对分离得到的菌株进行产胞外蛋白酶和纤维素酶活性测定和产胞外多糖能力检测。【结果】从西南地区4个湖泊中共分离筛选得到41株细菌,其中LB 15株、ZM 13株、SD 7株、TC 6株。根据16S r RNA基因序列的系统进化分析,4个地区可培养细菌的组成和丰度存在明显差异,其中LB和ZM的优势菌属是芽孢杆菌属(Bacillus),其次是气单胞菌属(Aeromonas)和假单胞菌属(Pseudomonas),分离的TC菌株全部属于芽孢杆菌属(Bacillus),分离的SD菌株特异性较强。进一步酶活性和胞外多糖检测表明,分离得到的41株细菌中有28株菌的发酵产物具有蛋白酶活性,6株具有纤维素酶活性,17株可产胞外多糖(Exopolysaccharides,EPS)。其中有2株细菌同时产蛋白酶、纤维素酶和胞外多糖,10株细菌同时产蛋白酶和胞外多糖,2株细菌同时产蛋白酶和纤维素酶,1株细菌同时产纤维素酶和胞外多糖。【结论】西南4个高山湖泊中存在丰富的微生物菌种资源,且4个湖泊中筛选的可培养细菌受所处环境的影响大。其中莲花湖由于高海拔和较偏僻等特点,人为干扰小,分离得到的细菌类群与其他湖泊相比明显不同;而马湖、凯邦亚湖和青海湖3个湖泊的海拔相对较低,受人类活动影响较大,分离得到的细菌均较常见。此外高山湖泊中的可培养细菌具有分泌多种胞外活性物质特性,为工业化应用奠定了资源基础,极具更深入的开发和研究价值。
张姜黄嘉丰李艳玲刘丹吴日帮廖斌强雷鸣肖潇武翠玲何海伦
关键词:高山湖泊RNA基因细菌多样性蛋白酶纤维素酶胞外多糖
鲮鱼皮胶原肽的制备及其抗氧化活性的检测被引量:4
2016年
为了以细菌胞外蛋白酶酶解低值蛋白资源制备抗氧化活性肽以及挖掘新型蛋白酶,采用液体发酵培养的方法对假交替单胞菌Pseudoalteromonas sp.SHK1-2进行发酵产酶,获得胞外蛋白酶粗酶液用于酶解热水法提取的鲮鱼胶原蛋白,通过超滤、sephadex LH-20分子筛层析获得具有DPPH自由基清除能力(35.6%±7%)、氧自由基清除能力(Oxygen radical absorbance capacity,ORAC)及DNA氧化损伤抑制活性的小分子肽,液相色谱质谱联用鉴定该活性肽分子量为776.2 Da,预测氨基酸序列为Thr-Ala-Gly-His-Pro-Gly-Thr-His。ORAC检测验证了人工合成的多肽同样具有抗氧化活性。研究表明细菌胞外蛋白酶在低值资源高值化中具有一定的应用前景,对于新型蛋白酶及其应用的挖掘具有一定的参考意义。
武翠玲吴日帮刘丹杨兴昊张姜黄嘉丰何海伦
关键词:抗氧化肽鲮鱼DPPHORACDNA氧化损伤
A rapid method for the detection of humic acid based on the poly(thymine)-templated copper nanoparticles
2018年
This work described a new method for the detection of humic acid(HA) based on the poly(thymine)(polyT)-templated copper nanoparticles(CuNPs). Without the presence of HA, the formation of poly Ttemplated CuNPs could take place, resulting in strong fluorescence emission peaks at 610 nm(upon excitation at 340 nm). On the other hand, when HA was present, strong interaction between HA and Cu^(2+)took place, which then hampered the effective formation of fluorescent CuNPs, leading to the decrease in fluorescence intensity. Furthermore, under the optimal experimental conditions, the method exhibited a high specificity to HA with a detection limit of 0.4 mg/L. This work has demonstrated a low-cost and convenient method that could be accomplished within 10 min. The method could provide a simple, rapid,and sensitive fluorescent platform for the detection of HA.
Changbei MaMingjian ChenHaisheng LiuKefeng WuHailun HeKemin Wang
Label-free fluorescent strategy for sensitive detection of tetracycline based on triple-helix molecular switch and G-quadruplex被引量:5
2017年
In this assay, a label-free fluorescent sensing platform based on triple-helix molecular switch(THMS) and G-quadruplex was developed for the detection of tetracycline. We demonstrated this approach by using THMS, which consists of a central section with a shortened 8-mer aptamer sequence with high affinity to tetracycline and flanked by two arm segments. G-rich oligonucleotide can specifically bind to thioflavin T(Th T) as a signal transduction probe(STP). In the absence of tetracycline, THMS remains stable, the fluorescence of background is low. By the addition of target tetracycline, the aptamer-target binding results in the formation of a structured aptamer-target complex, which disassembles the THMS and releases the STP. The free STP self-assembles into G-quadruplex and specifically binds to Th T which generates a obvious fluorescence enhancement. Using the triple-helix molecular switch, the developed aptamer-based fluorescent sensing platform showed a linear relationship with the concentration of tetracycline ranging from 0.2 to 20.0 nmol/L. The detection limit of tetracycline was determined to be970.0 pmol/L. The assay avoids complicated modifications or chemical labeling, making it simple and cost-effective. So, it is expected that this aptamer-based fluorescent assay could be extensively applied in the field of food safety inspection.
Tian-Xiao ChenFeng NingHai-Sheng LiuKe-Feng WuWei LiChang-Bei Ma
关键词:分子开关THMS
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