硝酸盐,镁盐对林可霉素生物合成的促进作用

在林肯链霉菌生物合成林可霉素代谢调节的研究中,发现硝酸盐可明显促进林可霉素的生物加入硝酸钾０．８％,林肯链霉菌合成林可霉素的产量可增加３７％．在发酵９６ｈ之前加入硝酸盐均能促进林可霉素的合成,但产量的增加随加入时间的延迟而降低。硝酸钾在促进产量的同时,使菌体生长减少,看来硝酸盐对林可霉素的合成与菌体生长之间起着调节作用。     

地中海诺卡氏菌(Nocardia mediterranei)合成力复霉素代谢调节的研究——Ⅰ.硝酸盐对地中海诺卡氏菌合成力复霉素SV的促进作用

本文报导了硝酸盐促进力复霉素生物合成现象的初步观察结果。加入硝酸钾0.8％,地中海诺卡氏菌(Nocardia mediterranei)NG 12—4合成力复霉素SV的产量可增加1.7倍。在发酵96小时之前加入硝酸盐均能促进力复霉素SV的合成,但产量的增加随加入时间的延迟而降低。硝酸钾在促进产量的同时,使菌体生长减少,看来硝酸盐对力复霉素SV的合成与菌体生长之间起着调节作用。 洗涤菌体试验指出,硝酸盐的加入诱导了力复霉素合成所需要的酶系,蛋白陈不能代替硝酸盐,进一步说明硝酸钾的作用并不是作为氮源利用。在蛋白质合成抑制剂氯霉素存在下,硝酸盐不再能促进力复霉素的合成,说明氯霉素抑制了硝酸盐所诱导的酶系的合成。 铵盐明显地抵消了硝酸盐对力复霉素合成的促进作用,可能是由于铵盐阻遏了硝酸还原酶的合成。 KNO~3~-菌体与-KNO~3~-菌体的形态有明显差别。两种菌体的脂肪含量也不相同。 KNO~3~-一菌体的脂肪含量为5.7％,而-KNO~3~-菌体则高达13.6％,看来硝酸盐在脂肪合成与力复霉素合成两条途径之间起着调节作用。     

林肯链霉菌合成林可霉素代谢调节的研究

在摇瓶条件下研究了葡萄糖、铵盐、磷酸盐对林可霉素产生菌林肯链霉菌的生长及林可霉素生物合成的影响。发酵过程中林可霉素的合成主要发生在菌体生长期,逐渐下降。使用6%的葡萄糖未发现通常所说的“葡萄糖效应”。0.2%铵盐有利于细胞生长,但0.8%NH+4对林可霉素的生物合成具有抑制作用。发酵48h后补加0.6% NH^＋_４,对林可霉素的生成没有显著影响。0.05%～0.1%磷酸盐对林可霉素合成具有较强的抑制作用。并就磷酸盐对菌体由初级代谢转向次级代谢的作用作了初步考察。     

林肯链霉菌谷氨酰胺合成酶活力调节的研究

对不同氮源生长条件下林肯链霉菌无细胞粗提液中谷氨酰胺合成酶 (GS)的研究结果表明 ,高浓度NH+4阻遏了GS的生物合成。从不同氮源生长条件下林肯链霉菌中分离纯化了GS ,其性质没有差别。以受腺苷化调节的产气克雷伯氏菌GS作对照 ,林肯链霉菌GS没有明显的氨休克作用 ,经蛇毒磷酸二酯酶处理后 ,其活力没有变化。这些结果都说明林肯链霉菌GS不存在腺苷化共价修饰这一调节方式。反馈抑制作用是林肯链霉菌GS的一种重要的调节方式 ,这种抑制作用是以累积的方式进行的 ,这表明各种抑制剂对GS作用位点不同 ,各种抑制剂对GS的抑制作用是相互独立的。由此推测 ,林肯链霉菌GS是一种变构酶。     

链霉菌形态分化与次级代谢产物合成的研究进展

链霉菌是一类具有复杂的形态分化周期和强大的次级代谢能力的高GC含量的放线菌,能够利用其初级代谢产生的前体化合物和能量,合成多种结构复杂、功能多样的具有生物活性的次级代谢产物,在农业、食品、畜牧业、工业以及医药研究等领域都具有重要的价值。在链霉菌的形态分化后期常常伴随着次级代谢产物的生物合成,并且两者都受到复杂的网络调控;同时链霉菌的形态对次级代谢产物的产量和种类造成很大影响。对链霉菌生长周期的全面理解将加深对链霉菌形态分化与次级代谢产物合成关系的认识。本文将对链霉菌的形态分化过程、形态分化和抗生素合成两者共同的调控因子以及链霉菌形态与抗生素产量之间的关系进行综述,这将有助于理解抗生素的合成过程,也将会在缩短发酵周期、构建高产工程菌株、新型杀菌剂的研发以及新型抗生素的合成等方面给予我们启发。     

甲壳素促进茂源链霉菌发酵产酶

为了提高茂源链霉菌发酵生产谷氨酰胺转胺酶的产量,研究了甲壳素对茂源链霉菌发酵产酶的影响。结果表明,添加0.5%的甲壳素对茂源链霉菌发酵产酶的促进效果极显著,但甲壳素的添加量达到2%时反而会抑制菌株产酶。从菌株生长代谢过程中p H变化、产酶情况、发酵液中蛋白含量及总氮含量等方面,对甲壳素促进茂源链霉菌发酵产酶的作用机理进行了初步探讨。研究显示,甲壳素在茂源链霉菌发酵过程中对菌体生长产生一定的胁迫,刺激菌体大量分泌次级代谢产物,从而提高茂源链霉菌的产酶。对菌株发酵过程的显微观察则表明,甲壳素也可能通过分散菌体生长,提高菌体向胞外分泌谷氨酰胺转胺酶的量来促进产酶。     

透明颤菌血红蛋白基因表达对金色链霉菌生长代谢的影响

利用四环素抗性基因启动子在金色链霉菌中表达透明颤菌血红蛋白基因。在1m3发酵罐中研究了工程菌株的生长代谢特性。在溶解氧充足的条件下,透明颤菌血红蛋白表达,对金色链霉菌生长代谢未产生明显影响,工程菌株与参比菌株的生长代谢特性基本一致,工程菌株和参比菌株金霉素最终浓度分别为22905u/mL、22896u/mL。在低溶解氧条件下,透明颤菌血红蛋白的表达,可促进金色链霉菌菌体生长、菌丝活力保持和金霉素的合成：工程菌菌体浓度比参比菌株高5％～10％,产物合成提高11.4％。     

信号分子在链霉菌天然产物发现和开发中的应用研究进展

链霉菌具有巨大的合成次级代谢产物的潜力,但在实验室常规培养条件下链霉菌中大部分生物合成基因簇是沉默的,或者表达量极低。链霉菌中信号分子可调节形态分化和代谢产物的生物合成。通过对编码这些信号分子合成酶或受体的基因进行操作,或在发酵液中添加信号分子,可以激活链霉菌中的沉默生物合成基因簇,发现新的天然产物,或者提升已发现的天然产物产量。本文以γ-丁内酯和γ-丁烯内酯两类信号分子为例总结了过去十余年中信号分子在链霉菌天然产物发现和产量提升中的应用,以期为微生物天然产物的开发提供借鉴。     

链霉菌中高效生产聚酮化合物的研究方法及进展北大核心CSCD

聚酮化合物是通过聚酮合成途径产生的一大类结构和生物活性多样的次级代谢产物,是链霉菌产生的主要次级代谢产物,具有重要的经济价值。为了在链霉菌中提高聚酮化合物产量,以满足工业生产需求,近年来,代谢工程的方法被广泛应用,例如,过表达合成途径中限速酶或途径特异性激活蛋白、强化前体供应、去除产物反馈抑制、合成基因簇异源表达等。本文将从代谢工程改造实例入手,全面综述链霉菌中聚酮化合物高效生物合成的研究方法及进展,并对利用合成生物学策略智能动态适配各个相关途径,进而提高该类化合物产量的研究思路进行展望。     

无机磷酸盐对链霉菌合成次级代谢产物的影响

链霉菌一个突出的特征是具有合成丰富的次级代谢产物的能力,许多次级代谢产物,如抗生素、免疫抑制剂、抗癌物质等在临床医药、水产养殖业等领域具有重要的应用价值。链霉菌次级代谢产物的合成常与环境中的营养因子有着密切的关系,在代谢水平上综述了无机磷酸盐对链霉菌合成次级代谢产物的影响,并在转录水平上阐释了双组分信号转导系统PhoR-PhoP的分子调控机制。PhoR-PhoP能够感应环境中的无机磷酸盐信号,当无机磷酸盐的浓度低于一定"阈值"时,PhoP蛋白作为主导的调控因子将抑制参与中心代谢和次级代谢等一系列基因的转录表达,减慢磷酸盐的消耗,并激活磷酸盐的摄取和转运系统及时补充胞内的磷酸盐,最终影响链霉菌次级代谢产物的合成和形态发育分化。     

Effects of phosphate, glucose, and ammonium on cell growth and lincomycin production by Streptomyces lincolnensis in chemically defined media

Cell growth and lincomycin production were measured in batch cultures of Streptomyces lincolnensis in chemically defined media. In these fermentations the specific rate of antibiotic production was maximal during growth and always declined at the end of the growth phase. It was found that both phosphate and ammonium salts, while required for cell growth, had negative effects on antibiotic production. By increasing the concentration of magnesium sulfate, it was possible to increase both the production rates and final titers of lincomycin. The mechanism for this effect was found to be the reduction of soluble phosphate in the medium through the precipitation of ammonium magnesium phosphate. Lincomycin production rates were not inhibited by glucose at concentrations of up to 30 g/L.     

The culture of plant cells with ammonium salts as the sole nitrogen source

Soybean cell suspension cultures grew on defined media with ammonium as the sole nitrogen source if Krebs cycle acids were added. Satisfactory growth was obtained with ammonium salts of citrate, malate, fumarate, or succinate, when compared with the regular medium containing nitrate and ammonium. Little or no growth occurred when ammonium salts of shikimate, tartrate, acetate, carbonate, or sulfate were used. The cells also grew well with l-glutamine as nitrogen source. The specific activities of glutamine synthetase and isocitrate dehydrogenase (nicotinamide adenine dinucleotide phosphate) were lower than in cells grown on a nitrate medium, but ammonium enhanced the activity of glutamate dehydrogenase. Cells of soybean, wheat, and flax have been cultured for an extended period on the ammonium citrate medium.     

Effect of nitrogen sources on cellulase biosynthesis by a mutant strain of Trichoderma viride 44

The effect of various nitrogen sources on cellulase biosynthesis by the mutant strain Trichoderma viride 44 was examined. This strain may utilized nitrogen in the nitrate, ammonium of organic form. When cultivating this strain, it appears advantageous to add to the nutrient medium yeast and yeast lyzates as well as their mixture with ammonium sulfate. Cellulase reached its maximum activity of 20.2, 21.5 and 23.2 mu/ml when grown on the medium containing ammonium phosphate, peptone and brewing yeast plus ammonium sulfate, respectively. It is useful to apply nitrogen in its organic forms in small quantities and in combination with mineral forms. The nitrogen presence in the medium is necessary only at the exponential stage of fungal growth. The lack of nitrogen in the stationary stage characterized by the maximum cellulase formation does not inhibit an increase in the enzyme activity.     

NUTRITIONAL REQUIREMENTS FOR OVULE FORMATION IN EXCISED PISTILS OF NIGELLA

The nutritional requirements for ovule formation in Nigella saliva L. were investigated by growing excised pistils on defined media. Pistils grown on a medium containing the minerals of Murashige and Skoog produced significantly more ovules than on a medium containing the minerals of Bilderback. When the nitrogen, sulfate, and phosphate of the Bilderback medium were adjusted to levels comparable to those of the Murashige and Skoog medium, a similar number of ovules was formed. The effect of different forms and concentrations of nitrogen on ovule formation and pistil growth was investigated. High concentrations of nitrate (40 mil) favored pistil growth and ovule formation, but comparable levels of ammonium were toxic. When ammonium at concentrations above 10 mM was added to nitrate media, ovule formation was inhibited. A medium containing low concentrations of ammonium (10 mM) and nitrate (5 mM) supported more ovule formation and pistil growth in young pistils than a low-nitrate (5 mM) medium without ammonium. However, ovule formation on a medium containing 10 mM ammonium and 5 mM nitrate was significantly less than on a medium containing only 15 mM nitrate. Low concentrations of organic nitrogen in the form of α-alanine (1 mM) and γ-aminobutyric acid (5 mM) supported ovule formation and pistil growth similar to a high nitrate medium.     

Competition between the green alga Scenedesmus and the cyanobacterium Synechococcus under different modes of inorganic nitrogen supply

In this study, we evaluated growth responses of the green alga Scenedesmus and the cyanobacterium Synechococcus supplied with inorganic nitrogen in different ways. A competitive situation in which nitrogen was limiting was created in mixed cultures as well as in cultures growing in the same vessel but separated by a permeable dialysis membrane. Supplying inorganic nitrogen in small pulses at a high frequency favoured the cyanobacterium Synechococcus, whereas batch additions favoured the green alga Scenedesmus. When using a large-pulse/low-frequency supply mode, the yield of the green alga was higher when ammonium was added as nitrogen source compared to when nitrate was added. By contrast, the yield of the cyanobacterium was higher in the nitrate regime. However, uptake experiments using unialgal cultures showed that both organisms depleted the medium of ammonium more rapidly than they depleted the medium of nitrate; i.e. the higher yield of the cyanobacterium in the nitrate regime than in the ammonium regime can be attributed to the effects of competition with the green alga. Since nitrate assimilation involves the consumption of reductive power, we suggest that the outcome of competition was governed by the fact that green alga was light limited and therefore better able to compete for ammonium than for nitrate. The results from the laboratory studies are discussed in relation to results from an enclosure experiment performed in Lake Erken, Sweden. In that field experiment, in which additions of both phosphate and ammonium were applied every second day to 350-l enclosures, the green algal biomass increased exponentially during an incubation period of 22 days.     

Removal of nitrate, nitrite, ammonium and phosphate ions from water by the aerial microalga Trentepohlia aurea

The aerial microalga Trentepohlia aurea has beeninvestigated in relation to removal characteristics of nitrate, nitrite,ammonium and phosphate ions. When the alga was cultured in medium with veryhighconcentrations of ammonium, nitrate and phosphate ions, it showed relativelyhigh growth and removal rates. It also grew quite well with high nitriteconcentration (< 141 mg NO₂-N L^–1).The removal rate was 0.28 mg NO₂-N L^–1day^–1 in the 40-day culture, when it was cultured in modifiedBold's basal medium with added 51 mg NO₂-NL^–1. In addition, we examined simultaneous removal of nutrientions. The biomass was 1.5 times higher in medium which N- and P-sourcesufficient than in ordinary medium. Higher removal ratios of nitrite andnitratefrom medium were shown in a 30-day culture, reaching 37% and 32%, respectively.It is concluded that T. aurea has the potential for use inthe purification of wastewater.     

The action of different biocides on the phosphatase activity of Aspergillus niger

Metalorganic and quarternary ammonium compounds when added to culture medium inhibited growth of Aspergillus niger mycelium and activity of neutral and alkaline phosphatase. A quarternary ammonium compound, ethonium, and a tin-organic compound, tributyl oxide, exerted an inhibiting effect on activity of acid phosphatase which amounted to 54% of the total phosphatase activity in mycelium and 94% in the culture liquid. The rest of biocides induced lysis of intracellular membranes, phosphatase release from lysosomas, which made acid phosphatase activity higher. Being introduced into the mycelium homogenate the above compounds inhibited activity of the acid phosphatase. The same biocides inhibited extracellular acid phosphatase in the culture liquid. Recommendations are given on the use of a number of substances as means for protection of industrial materials from biolesions.     

Effect of different forms of nitrogen in the biosynthesis of gentamicin by a Micromonospora purpurea var. violacea 1935 culture]

The effect of different inorganic sources of reduced and oxidized nitrogen on biosynthesis of gentamicin was studied. It was shown that both the ammonium and the nitrate nitrogen were consumed by the antibiotic-producing organism. Out of all the salts tested only ammonium sulfate stimulated the biosynthesis of gentamicin. The positive role of this salt was due to both the ammonium and the sulfogroup, since the presence of the sulfogroup alone in sodium sulfat, magnesium or sulfuric acid resulted only in partial stimulation of gentamicin biosynthesis. Simultaneous addition of sulfuric acid ammonium nitrate which suppressed the antibiotic biosynthesis when used alone was equal to introduction of ammonium sulfate.     

Effect of glycerin on the biosynthesis of heliomycin by Streptomyces olivocinereus

Glycerol as the sole carbon source was added to the medium or biosynthesis of heliomycin by Streptomyces olivocinereus and the effect of its concentration on the culture growth and antibiotic production was studied. The culture growth and the amount of the antibiotic synthesized per 1 unit of the fermentation broth were limited by glycerol added in quantities of 0.05 to 1 per cent. Further increasing of the glycerol concentration had no significant effect on the culture growth and antibiotic biosynthesis. The amount of the antibiotic synthesized per 1 unit of the mycelial mass relatively slightly depended on the glycerol concentration. The rate of glycerol consumption by the young 24-hour culture in batch fermentations markedly exceeded that of glycerol consumption by the 48-hour culture. The younger mycelium significantly increased its rate of glycerol consumption when the initial concentration was increased whereas the rate of glycerol consumption by the more mature mycelium did not depend on the initial concentration of the carbon source (within 0.5-2 per cent). The rate of heliomycin biosynthesis practically slightly depended on the initial concentration of glycerol.