DOPA Production with Enterobacter cloacae NB 320 by Transamination Reaction

Taxonomical investigation was performed on the bacterium, strain NB 320 isolated from soil, and it was identified as Enterobacter cloacae. This bacterium produced the enzyme which catalyzed the transamination reaction between 3,4-dihydroxyphenyl pyruvate and an amino acid to form l-Dopa.

The optimum culture conditions for the enzyme production were studied along with the characteristics of the enzyme. The enzyme of the strain was different in some properties from that of Alcaligenes faecalis IAM 1015 which had been already studied. The former utilized glutamate as an amino donor best among the amino acids tested for transamination and was induced by the addition of glutamine and asparagine. Intact cells of the strain did not catalyze the reaction unless they were treated with sonication or with a detergent.     

New Antibacterial Diterpenoids from the Sarcodon scabrosus Fungus

The structures of new antibacterial diterpenoids that had been isolated from Sarcodon scabrosus were established by chemical and spectral means to be sarcodonin L (2) and M (3), both having the cyathane skeleton. Other antibacterial compounds were identified to be allocyathin B₂ (1), sarcodonin G (4) and sarcodonin A (5) by comparing their spectral data with those of authentic samples.     

Distribution and Solubilization of Particulate Gluconate Dehydrogenase and Particulate 2-Ketogluconate Dehydrogenase in Acetic Acid Bacteria

The distribution of two particulate enzymes, gluconate dehydrogenase (GDH) and 2-ketogluconate dehydrogenase (2KGDH), was investigated with cell free extract through 26 strains of genus Acetobacter and genus Gluconobacter. GDH activity was found in the cell free extracts from all strains of genus Gluconobacter and two species of genus Acetobacter, A. aceti and A. aurantium. High activity of 2KGDH was also found in the pigment-producing strains of genus Gluconobacter.

Best solubilization of particulate enzymes was attained with the highest recovery when 10 mg of Triton X–100 and 30 mg of protein of particulate fractions in 1 ml of 0.01 m phosphate buffer, pH 6.0, are incubated for 9 hr at 5°C with continuous stirring.

By comparison of the total enzyme activity of particulate enzymes with that of NAD(P)-linked enzymes in the cell free extract, it was obvious that the formation of ketogluconates by particulate enzymes was much more predominant, roughly over 100 times higher, as that of NAD(P)-linked enzymes.     

Studies on γ Globulin of Rice Embryo

An improved method has been described for the isolation and purification of γ globulin from rice embryo. The method involves the extraction with phosphate buffer, pH 7.0 and ionic strength 0.1, the fractionation in saline solution of ionic strength 0.31, the removal of nucleic acids by precipitation with ammonium sulfate and the gel filtration chromatography on a Sephadex G-200 column. Although the preparations exhibited homogeneous patterns in sedimentation analysis, the electrophoretic patterns on polyacrylamide gels at pH 8.35 and ionic strength 0.11 exhibited at least two components. Three major components, γ₁, γ₂ and γ₃ globulins, were isolated by ion exchange chromatography on a DEAE Sephadex A-50 column. These components were revealed to be homogeneous in electrophoresis as well as sedimentation. N-Terminal amino acid compositions have also been described.

The molecular weight of γ₁ globulin was determined as 2.0 × 10⁵ by the Archibald method, and the intrinsic viscosity, [η], and the sedimentation coefficient, s_{20, w}^°, were found to be 0.0424 dl/g and 7.26S respectively. These values indicated the large asymmetry of the protein. The protein was composed of 18 residues of hexose, 3 residues of pentose, 6 residues of hexosamine and 1751 residues of amino acids: Lys₅₈, His₄₇, Arg₁₄₈, Asp₁₂₆, Glu₂₇₃, Gly₁₆₁, Ala₁₄₄, Val₁₂₁, Leu₁₀₆, Ile₇₂, Pro₈₃, Ser₁₃₆, Thr₄₈, Hyp₆₈, Cys₁₇, Met₁₆, Tyr₄₄, Trp₈, Phe₇₅ and amide ammonia₁₆₃. The N-terminal amino acid analysis suggested that the protein was composed of ten subunits. The properties and the composition were discussed in comparison with those of the 7S globulin of soybean cotyledon.     

马特链霉菌7-木糖紫杉烷糖基水解酶初步研究

酶法转化7-木糖紫杉烷(7-XDT)为10-脱乙酰基紫杉醇(10-DAT)是目前合成紫杉醇的最主要途径.本研究分离到一株具有产生7-木糖紫杉烷(7-XDT)糖基水解酶能力的马特链霉菌(Streptomyces matensi YUCM 410051),通过酶的最适反应温度、硫酸铵分级沉淀、最适反应pH和酶的有机试剂耐受等研究,发现最适反应温度为25～30℃,硫酸铵分级沉淀酶活在20％～70％的盐浓度时活性最高;粗酶液最适反应pH在6.0～7.5,酶的甲醇耐受浓度和DMSO耐受浓度均为10％.深入研究该酶对开发具有水解7-木糖紫杉烷(7-XDT)中木糖基的酶资源和提高红豆杉中的紫杉烷类化合物的利用率具有重要价值.     

北部湾近海珊瑚礁区系沉积物抗菌活性放线菌类群

通过对大肠埃希菌和枯草芽胞杆菌抗菌活性初步筛选,从北部湾近海珊瑚礁区5个沉积物样品中成功分离得到51株具有不同抗菌活性的放线菌,其中9株具有较强抗菌能力。根据这9株放线菌的菌落和孢子形态,可确定它们都属于链霉菌属。 RAPD-PCR分析表明这9株放线菌为6种不同类型,16S rDNA序列和系统发生树分析表明,9株放线菌可划分到4个大的类群6种不同类型,且结果显示RAPD-PCR聚类分析与16S rDNA序列聚类分析的结果具有较大的一致性。生理生化鉴定结果表明,分离株与亲缘关系最近的放线菌模式菌株的生理生化特征均存在差异,这说明分离株为放线菌新种的可能性比较大。这6种放线菌具有较为广谱的抑菌活性,并且抑菌活性均存在一定的差异,说明其可能分泌出多种结构功能不同的活性次生代谢产物。研究结果表明,广西北部湾近海珊瑚礁区系沉积物蕴藏着丰富的可供药物开发的放线菌资源。     

南海东海岛沉积物分离细菌的鉴定及其抑菌活性

[背景] 海洋微生物在活性物质开发方面具有巨大的应用前景,而目前有关南海东海岛微生物的研究鲜少。[方法] 对从东海岛沉积物中分离纯化的海洋细菌,采用形态学观察、生理生化以及16S rRNA基因序列的系统发育分析方法进行鉴定;以大肠杆菌（Escherichia coli）、枯草芽孢杆菌（Bacillus subtilis）和金黄色葡萄球菌（Staphylococcus aureus）作为指示菌,测定其抑菌活性;对具有抑菌活性的菌株扩增聚酮合酶（Polyketide synthase I,PKSI）基因,并与已知的PKSI氨基酸序列比对;选择具有PKSI基因的代表菌株,检测菌株及其发酵抑菌物的稳定性。[结果] 分离纯化到25株海洋细菌,分属于不动杆菌属（Acinetobacter）、交替单胞菌属（Alteromonas）、芽孢杆菌属（Bacillus）、嗜冷杆菌属（Psychrobacter）、假交替单胞菌属（Pseudo-alteromonas）、海洋单胞菌属（Oceanimonas）、葡萄球菌属（Staphylococcus）、微球菌属（Micrococcus）和海杆菌属（Marinobacter）。12株菌株通过基因筛选检测到PKSI编码基因,其中6株菌株具有抑菌活性和PKSI编码基因,并分属于芽孢杆菌属和交替单胞菌属;PKSI氨基酸序列同源性分析推测菌株DHD-15和DHD-a可能产生新的I型聚酮合酶结构。菌株DHD-15和DHD-L生长温度范围为15-40℃,可耐受10% NaCl高盐以及pH 3和pH 11的酸碱条件,但不耐高温;菌株DHD-15产生的抑菌物质可耐受100℃和pH 11的高温碱性条件,在50℃、pH 9条件下制备和室温保藏条件下抑菌活性较高,其稳定性较好。[结论] 南海东海岛沉积物筛选的细菌种具有抑菌活性,具有产聚酮类活性物质的潜力。     

TBK1 directs WIPI2 against Salmonella

Defense of the mammalian cell cytosol against Salmonella invasion is reliant upon capture of the infiltrating bacteria by macroautophagy (hereafter autophagy), a process controlled by the kinase TBK1. In our recent study we showed that recruitment of TBK1 activity to Salmonella stabilizes the key autophagy regulator WIPI2 on those bacteria, a novel and essential function for TBK1 in the control of the early steps of antibacterial autophagy. Substantial redundancy exists in the precise recruitment mechanism for TBK1 because engagement with any of several Salmonella-associated ‘eat-me’ signals, including host-derived glycans, and K48- and K63-linked ubiquitin chains, suffices to recruit TBK1 functionality. We therefore propose that buffering TBK1 recruitment against potential bacterial interference might be of evolutionary advantage to the host.     

羊毛硫肽类化合物(Lanthipeptide)生物合成新进展

羊毛硫肽化合物(Lanthipeptides)是由核糖体合成并经过翻译后修饰得到的一大类肽类天然产物。这类化合物广泛的产生于不同种类的细菌,具有丰富的结构和生物活性多样性,为活性药物研究和开发提供重要的来源。本文综述了近几年来羊毛硫肽化合物生物合成进展,从其合成酶结构,进化机制,区域和立体选择性控制等方面进行了简要的讨论,展示了羊毛硫肽类化合物生物合成中特殊而迷人的酶学机制。     

靶向铜绿假单胞菌(Pseudomonas aeruginosa)粘肽合成酶PBP3的抗菌先导化合物的虚拟筛选及活性研究

【目的】开发出与铜绿假单胞菌粘肽合成酶PBP3有高亲和力,具有全新结构的先导化合物。【方法】以铜绿假单胞菌粘肽合成酶PBP3为靶点,通过分子对接软件DOCK6.5,对含有104万个小分子化合物的数据库进行了大规模虚拟筛选,选取结构相对简单的中选化合物进行合成,得到先导化合物,并验证其抑菌活性。【结果】通过grid score进行第一轮初筛,筛选出grid score分值小于–30 kcal/mol的6万个化合物,接着以amber score进行第二轮筛选,筛出amber score分值小于–20 kcal/mol的化合物约200个。最终,经过观察分析,从中挑选出4种打分高并且结构新颖的小分子化合物作为先导化合物。合成出的先导化合物及其衍生物对铜绿假单胞菌等常见微生物的最小抑菌浓度(MIC)在175–275μg/m L之间,对革兰氏阴性菌和阳性菌均有效,MIC值比作为阳性对照的磺胺嘧啶更低,说明先导化合物具有较好的抗菌活性。【结论】这些先导化合物可以进一步开发为新型抗菌药物,用于解决铜绿假单胞菌的耐药性问题。