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Carbohydrate Metabolism in Streptomycetes II. Isolation and Enzymatic Synthesis of Trehalose 总被引:1,自引:0,他引:1 下载免费PDF全文
Alan D. Elbein 《Journal of bacteriology》1967,94(5):1520-1524
A number of streptomycetes were examined for their ability to synthesize trehalose phosphate as well as for the presence of α,α-trehalose. In each case, an enzyme system was demonstrated which catalyzed the transfer of glucose from guanosine diphosphate-glucose to glucose-6-phosphate to form trehalose phosphate. Thus, this group of organisms appears to synthesize trehalose phosphate by a different mechanism from that described in insects, yeast, and fungi. In addition, trehalose was isolated from each of these organisms. In several of these cases, crystallization of the sugar and determination of the physical properties showed that the sugar was α,α-trehalose. 相似文献
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Trehalose is a non-reducing disaccharide, which can protect proteins, lipid membranes, and cells from desiccation, refrigeration, dehydration, and other harsh environments. Trehalose can be produced by different pathways and trehalose synthase pathway is a convenient, practical, and low-cost pathway for the industrial production of trehalose. In this study, 3 candidate treS genes were screened from genomic databases of Pseudomonas and expressed in Escherichia coli. One of them from P. stutzeri A1501 exhibited the best transformation ability from maltose into trehalose and the least byproduct. Thus, whole cells of this recombinant E. coli were used as biocatalyst for trehalose production. In order to improve the conversion rate of maltose to trehalose, optimization of the permeabilization and biotransformation were carried out. Under optimal conditions, 92.2 g/l trehalose was produced with a high productivity of 23.1 g/(l h). No increase of glucose was detected during the whole course. The biocatalytic process developed in this study might serve as a candidate for the large scale production of trehalose. 相似文献
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以大米淀粉为原料,多酶复配制备海藻糖。确定了实验室条件下多酶复配生产海藻糖的最佳条件:以15%(m/V)大米淀粉为底物,催化温度45℃、pH 6. 0、DE值16、α/β-CGTase加量为1. 4U/ml、催化28h后糖化处理12h,海藻糖转化率由双酶法催化的50%提高至73%。在底物浓度为25%(m/V)时,海藻糖产量最高达到182. 5g/L,随后对高浓度海藻糖进行分离提取,分别考察了活性炭脱色、离交分离、浓缩结晶等对海藻糖提取效率的响。 相似文献
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Trehalose Toxicity in Cuscuta reflexa: Cell Wall Synthesis Is Inhibited upon Trehalose Feeding 下载免费PDF全文
α,α-Trehalose induced a rapid blackening of the terminal 2.5-centimeter region of excised Cuscuta reflexa Roxb. vine. The incorporation of radioactivity from [14C]glucose into alkali-insoluble fraction of shoot tip was markedly inhibited by 12 hours of trehalose feeding to an excised vine. This inhibition was confined to the apical segment of the vine in which cell elongation occurred. The rate of blackening of shoot tip explants was hastened by the addition of gibberellic acid A3, which promoted elongation growth of isolated Cuscuta shoot tips. The symptom of trehalose toxicity was duplicated by 2-deoxyglucose, which has been shown to be a potent inhibitor of cell wall synthesis in yeast. The observations suggest that trehalose interferes with the synthesis of cell wall polysaccharides, the chief component of which was presumed to be cellulose. 相似文献
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Trehalose phosphorylase from Euglena gracilis 总被引:2,自引:0,他引:2
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将B. circulans 251 β-CGTase应用于海藻糖制备,海藻糖转化率从50.4%提高至71.9%。为进一步提高底物的转化率,运用易错PCR-高通量筛选技术筛选对以麦芽糖为歧化反应受体的亲和性提高的B. circulans 251 β-CGTase突变体。利用低底物浓度的96孔板4,6-亚乙基-对硝基苯-α-D-麦芽七糖苷(EPS)显色法,最终筛选得到了一株对麦芽糖亲和性提高的突变体M234I。将野生型β-CGTase和突变体酶M234I进行蛋白质纯化,测定其酶学性质。结果表明,突变体的比活为345.25U/mg,野生型则为357.63U/mg;突变体M234I对麦芽糖的Km为0.258 2mmol/L,仅为野生型(0.474 9mmol/L)的54.4%,对麦芽糖的亲和性显著提高;突变体的最适温度、最适pH较野生型未发生较大变化。以麦芽糊精(DE值16)为底物,将突变体M234I用于多酶复配体系生产海藻糖,酶反应结果表明海藻糖的转化率最高达74.9%,较野生型β-CGTase提高约3%。 相似文献
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Trehalose Synthesis by Sequential Reactions of Recombinant Maltooligosyltrehalose Synthase and Maltooligosyltrehalose Trehalohydrolase from Brevibacterium helvolum 总被引:3,自引:0,他引:3 下载免费PDF全文
Yong Hwan Kim Tae Keun Kwon Sungsoon Park Hak Soo Seo Jong-Joo Cheong Chung Ho Kim Ju-Kon Kim Jong Seob Lee Yang Do Choi 《Applied microbiology》2000,66(11):4620-4624
A DNA fragment encoding two enzymes leading to trehalose biosynthesis, maltooligosyltrehalose synthase (BvMTS) and maltooligosyltrehalose trehalohydrolase (BvMTH), was cloned from the nonpathogenic bacterium Brevibacterium helvolum. The open reading frames for the two proteins are 2,331 and 1,770 bp long, respectively, and overlap by four nucleotides. Recombinant BvMTS, BvMTH, and fusion gene BvMTSH, constructed by insertion of an adenylate in the overlapping region, were expressed in Escherichia coli. Purified BvMTS protein catalyzed conversion of maltopentaose to maltotriosyltrehalose, which was further hydrolyzed by BvMTH protein to produce trehalose and maltotriose. The enzymes shortened maltooligosaccharides by two glucose units per cycle of sequential reactions and released trehalose. Maltotriose and maltose were not catalyzed further and thus remained in the reaction mixtures depending on whether the substrates had an odd or even number of glucose units. The bifunctional in-frame fusion enzyme, BvMTSH, catalyzed the sequential reactions more efficiently than an equimolar mixture of the two individual enzymes did, presumably due to a proximity effect on the catalytic sites of the enzymes. The recombinant enzymes produced trehalose from soluble starch, an abundant natural source for trehalose production. Addition of α-amylase to the enzyme reaction mixture dramatically increased trehalose production by partial hydrolysis of the starch to provide more reducing ends accessible to the BvMTS catalytic sites. 相似文献
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《Bioscience, biotechnology, and biochemistry》2013,77(7):1328-1329
The production of ethanol from maltose by Zymobacter palmae T109 in monoculture fermentations, and in co-culture fermentations together with Zymomonas mobilis B69 was studies. Zymobacter palmae T109, produced 5.5% (w/v) of ethanol when co-cultured with Zymomonas mobilis B69, but Zymobacter palmae T109 produced only 4.9% (w/v) ethanol from 15% (w/v) maltose medium in monoculture fermentation. 相似文献
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Atsumi Kamogawa Kozi Yokobayashi Toshio Fukui 《Bioscience, biotechnology, and biochemistry》2013,77(12):2813-2819
Maltose phosphorylase (EC 2.4.1.8) from Lactobacillus brevis was purified 29-fold over the crude extract. The final preparation was at least 80% pure and had a specific activity of 18 units/mg protein. The molecular weights of the native enzyme and of the component dissociated in sodium dodecyl sulfate were 150,000 and 80,000, respectively. The enzyme does not contain pyridoxal-5′-phosphate as a cofactor. It can not act on maltitol, malto-triitol, sucrose, lactose and trehalose, and essentially not on isomaltose, maltobionic acid, maltotriose and maltotetraose. Inhibitory effect was observed with CuSO4, HgCl2 and p-chloromercuribenzoate. Some other properties were also examined. A possibility of using this enzyme for the analysis of maltose was proposed. 相似文献
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Geigert J Neidleman SL Dalietos DJ Dewitt SK 《Applied and environmental microbiology》1983,45(2):366-374
The enzymatic synthesis of alpha,beta-halohydrins from gaseous alkenes is described. The enzymatic reaction required an alkene, a halide ion, dilute hydrogen peroxide, and a haloperoxidase enzyme. A wide range of gaseous alkenes were suitable for this reaction, including those containing isolated, conjugated, and cumulative carbon-carbon double bonds. Chlorohydrins, bromohydrins, and iodohydrins could be formed. The combining of this enzymatic synthesis with a previously described enzymatic synthesis of epoxides from alpha,beta-halohydrins provides an alternate pathway, other than the well-known enzymatic direct epoxidation pathway, from alkene to an epoxide. 相似文献
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Three Enzymes for Trehalose Synthesis in Bradyrhizobium Cultured Bacteria and in Bacteroids from Soybean Nodules 下载免费PDF全文
α,α-Trehalose is a disaccharide accumulated by many microorganisms, including rhizobia, and a common role for trehalose is protection of membrane and protein structure during periods of stress, such as desiccation. Cultured Bradyrhizobium japonicum and B. elkanii were found to have three enzymes for trehalose synthesis: trehalose synthase (TS), maltooligosyltrehalose synthase (MOTS), and trehalose-6-phosphate synthetase. The activity level of the latter enzyme was much higher than those of the other two in cultured bacteria, but the reverse was true in bacteroids from nodules. Although TS was the dominant enzyme in bacteroids, the source of maltose, the substrate for TS, is not clear; i.e., the maltose concentration in nodules was very low and no maltose was formed by bacteroid protein preparations from maltooligosaccharides. Because bacteroid protein preparations contained high trehalase activity, it was imperative to inhibit this enzyme in studies of TS and MOTS in bacteroids. Validamycin A, a commonly used trehalase inhibitor, was found to also inhibit TS and MOTS, and other trehalase inhibitors, such as trehazolin, must be used in studies of these enzymes in nodules. The results of a survey of five other species of rhizobia indicated that most species sampled had only one major mechanism for trehalose synthesis. The presence of three totally independent mechanisms for the synthesis of trehalose by Bradyrhizobium species suggests that this disaccharide is important in the function of this organism both in the free-living state and in symbiosis. 相似文献
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脂肪酶催化合成生物柴油的研究 总被引:78,自引:0,他引:78
生物柴油是用动植物油脂或长链脂肪酸与甲醇等低碳醇合成的脂肪酸甲酯,是一种替代能源。这里探讨了生物法制备生物柴油的过程,采用脂肪酶酯化和酯交换两条工艺路线进行催化合成。深入研究制备过程中,不同脂肪酶、酶的用量和纯度、有机溶剂、低碳醇的抑制作用、吸水剂的作用、反应时间和进程、底物的特异性和底物摩尔比等参数对酯化过程的影响。试验结果表明,采用最佳酯化反应参数和分批加入甲醇并用硅胶作脱水剂的工艺过程,酯化率可以达到92%,经分离纯化后的产品GC分析的纯度可达98%以上,固定化酶的使用半衰期可达到360h。同时对酯交换制备生物柴油过程中,甲醇的用量和甲醇的加入方式对脂肪酶催化过程的影响作了初步研究,优化后的酯交换率可达到83%。 相似文献
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Archaeal Binding Protein-Dependent ABC Transporter: Molecular and Biochemical Analysis of the Trehalose/Maltose Transport System of the Hyperthermophilic Archaeon Thermococcus litoralis 总被引:1,自引:0,他引:1 下载免费PDF全文
Reinhold Horlacher Karina B. Xavier Helena Santos Jocelyne DiRuggiero Marina Kossmann Winfried Boos 《Journal of bacteriology》1998,180(3):680-689
We report the cloning and sequencing of a gene cluster encoding a maltose/trehalose transport system of the hyperthermophilic archaeon Thermococcus litoralis that is homologous to the malEFG cluster encoding the Escherichia coli maltose transport system. The deduced amino acid sequence of the malE product, the trehalose/maltose-binding protein (TMBP), shows at its N terminus a signal sequence typical for bacterial secreted proteins containing a glyceride lipid modification at the N-terminal cysteine. The T. litoralis malE gene was expressed in E. coli under control of an inducible promoter with and without its natural signal sequence. In addition, in one construct the endogenous signal sequence was replaced by the E. coli MalE signal sequence. The secreted, soluble recombinant protein was analyzed for its binding activity towards trehalose and maltose. The protein bound both sugars at 85°C with a Kd of 0.16 μM. Antibodies raised against the recombinant soluble TMBP recognized the detergent-soluble TMBP isolated from T. litoralis membranes as well as the products from all other DNA constructs expressed in E. coli. Transmembrane segments 1 and 2 as well as the N-terminal portion of the large periplasmic loop of the E. coli MalF protein are missing in the T. litoralis MalF. MalG is homologous throughout the entire sequence, including the six transmembrane segments. The conserved EAA loop is present in both proteins. The strong homology found between the components of this archaeal transport system and the bacterial systems is evidence for the evolutionary conservation of the binding protein-dependent ABC transport systems in these two phylogenetic branches. 相似文献