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71.
Molecular architecture of DesV from Streptomyces venezuelae: a PLP-dependent transaminase involved in the biosynthesis of the unusual sugar desosamine 下载免费PDF全文
Burgie ES Thoden JB Holden HM 《Protein science : a publication of the Protein Society》2007,16(5):887-896
Desosamine is a 3-(dimethylamino)-3,4,6-trideoxyhexose found in certain macrolide antibiotics such as the commonly prescribed erythromycin. Six enzymes are required for its biosynthesis in Streptomyces venezuelae. The focus of this article is DesV, which catalyzes the PLP-dependent replacement of a 3-keto group with an amino functionality in the fifth step of the pathway. For this study the three-dimensional structures of both the internal aldimine and the ketimine intermediate with glutamate were determined to 2.05 A resolution. DesV is a homodimer with each subunit containing 12 alpha-helical regions and 12 beta-strands that together form three layers of sheet. The structure of the internal aldimine demonstrates that the PLP-cofactor is held in place by residues contributed from both subunits (Asp 164 and Gln 167 from Subunit I and Tyr 221 and Asn 235 from Subunit II). When the ketimine intermediate is present in the active site, the loop defined by Gln 225 to Ser 228 from Subunit II closes down upon the active site. The structure of DesV is similar to another sugar-modifying enzyme referred to as PseC. This enzyme is involved in the biosynthesis of pseudaminic acid, which is a sialic acid-like nonulosonate found in the flagellin of Helicobacter pylori. In the case of PseC, however, the amino group is transferred to the C-4 rather than the C-3 position. Details concerning the structural analysis of DesV and a comparison of its molecular architecture to that of PseC are presented. 相似文献
72.
近年来由多种致病链霉菌引起的马铃薯疮痂病在我国普遍流行,且危害程度逐年加重,严重影响块茎的品质和商品价值。病原菌土传和种传,难以防控。利用拮抗微生物抑制病菌生长是目前防控疮痂病的重要措施。【目的】从病薯田土样中定向筛选对马铃薯疮痂病具有显著防效的菌种,研究其拮抗机制,评价其环境适应性,为开发可产业化应用的高效复合功能菌剂提供理论依据。【方法】通过平板对峙及盆栽试验研究目标菌株对主要病原菌疮痂链霉菌Streptomyces scabies的抑制效果;采用形态学、生理生化实验及分子生物学方法,确定其分类地位;结合高效液相色谱质谱联用方法分析相关抑菌活性物质。【结果】获得3株对致病链霉菌S. scabies具有显著拮抗功能的菌株HZ11-4、HS-12、HZ13-1,抑菌圈直径分别为34、29、30 mm,对马铃薯微型薯疮痂病的防效分别为68.57%、57.15%和65.96%。菌体革兰氏染色呈阳性,经鉴定均为解淀粉芽孢杆菌Bacillus amylolique-faciens;3株菌皆可扩增出surfactin、iturin和fengycin等脂肽类物质合成酶相关基因片段,检测到上述脂肽类... 相似文献
73.
【背景】土霉素属四环素类抗生素,在医疗制药和动物养殖行业应用广泛。抗生素的滥用与残留极易导致环境中的益生菌产生耐药,各类益生菌通过食物进入机体后,可将耐药基因转移给致病菌。目前的耐药菌株筛选方法耗时较长,最小抑菌浓度测定结果也可能存在偏差,而且无法在抗菌机理及耐药机制方面提供线索。【目的】采用表面增强拉曼光谱(surface enhanced Raman spectroscopy,SERS)技术探究土霉素与植物乳杆菌之间的相互作用,以期为微生物耐药菌株的快速筛选提供理论基础,并为研究抗菌机理及耐药机制提供思路及方法。【方法】组装银纳米棒与石英后得到一种具有良好、稳定的SERS增强效应的固相基底;在植物乳杆菌菌液中加入不同浓度土霉素后各自分别孵育30、60和90 min,利用组装好的固相基底对细菌细胞进行SERS信号测定,拉曼光谱的出峰位置和峰强度变化可以体现不同土霉素浓度及作用时间条件下植物乳杆菌细胞成分的差异,可进一步推断土霉素对植物乳杆菌生长的影响。【结果】0.5×MIC土霉素会造成SERS光谱强度的降低;1.0×MIC土霉素作用90 min后,SERS在1 612、1 630 c... 相似文献
74.
Clara Torres-Barceló Mila Kojadinovic Richard Moxon R. Craig MacLean 《Proceedings. Biological sciences / The Royal Society》2015,282(1816)
Exposure to antibiotics induces the expression of mutagenic bacterial stress–response pathways, but the evolutionary benefits of these responses remain unclear. One possibility is that stress–response pathways provide a short-term advantage by protecting bacteria against the toxic effects of antibiotics. Second, it is possible that stress-induced mutagenesis provides a long-term advantage by accelerating the evolution of resistance. Here, we directly measure the contribution of the Pseudomonas aeruginosa SOS pathway to bacterial fitness and evolvability in the presence of sublethal doses of ciprofloxacin. Using short-term competition experiments, we demonstrate that the SOS pathway increases competitive fitness in the presence of ciprofloxacin. Continued exposure to ciprofloxacin results in the rapid evolution of increased fitness and antibiotic resistance, but we find no evidence that SOS-induced mutagenesis accelerates the rate of adaptation to ciprofloxacin during a 200 generation selection experiment. Intriguingly, we find that the expression of the SOS pathway decreases during adaptation to ciprofloxacin, and this helps to explain why this pathway does not increase long-term evolvability. Furthermore, we argue that the SOS pathway fails to accelerate adaptation to ciprofloxacin because the modest increase in the mutation rate associated with SOS mutagenesis is offset by a decrease in the effective strength of selection for increased resistance at a population level. Our findings suggest that the primary evolutionary benefit of the SOS response is to increase bacterial competitive ability, and that stress-induced mutagenesis is an unwanted side effect, and not a selected attribute, of this pathway. 相似文献
75.
F-type ATP synthases are rotary nanomotor enzymes involved in cellular energy metabolism in eukaryotes and eubacteria. The ATP synthase from Gram-positive and -negative model bacteria can be autoinhibited by the C-terminal domain of its ϵ subunit (ϵCTD), but the importance of ϵ inhibition in vivo is unclear. Functional rotation is thought to be blocked by insertion of the latter half of the ϵCTD into the central cavity of the catalytic complex (F1). In the inhibited state of the Escherichia coli enzyme, the final segment of ϵCTD is deeply buried but has few specific interactions with other subunits. This region of the ϵCTD is variable or absent in other bacteria that exhibit strong ϵ-inhibition in vitro. Here, genetically deleting the last five residues of the ϵCTD (ϵΔ5) caused a greater defect in respiratory growth than did the complete absence of the ϵCTD. Isolated membranes with ϵΔ5 generated proton-motive force by respiration as effectively as with wild-type ϵ but showed a nearly 3-fold decrease in ATP synthesis rate. In contrast, the ϵΔ5 truncation did not change the intrinsic rate of ATP hydrolysis with membranes. Further, the ϵΔ5 subunit retained high affinity for isolated F1 but reduced the maximal inhibition of F1-ATPase by ϵ from >90% to ∼20%. The results suggest that the ϵCTD has distinct regulatory interactions with F1 when rotary catalysis operates in opposite directions for the hydrolysis or synthesis of ATP. 相似文献
76.
Lin Wei Jiuxiang Gao Shumin Zhang Sijin Wu Zeping Xie Guiying Ling Yi-Qun Kuang Yongliang Yang Haining Yu Yipeng Wang 《The Journal of biological chemistry》2015,290(27):16633-16652
Cathelicidins are a family of gene-encoded peptide effectors of innate immunity found exclusively in vertebrates. They play pivotal roles in host immune defense against microbial invasions. Dozens of cathelicidins have been identified from several vertebrate species. However, no cathelicidin from marine reptiles has been characterized previously. Here we report the identification and characterization of a novel cathelicidin (Hc-CATH) from the sea snake Hydrophis cyanocinctus. Hc-CATH is composed of 30 amino acids, and the sequence is KFFKRLLKSVRRAVKKFRKKPRLIGLSTLL. Circular dichroism spectroscopy and structure modeling analysis indicated that Hc-CATH mainly assumes an amphipathic α-helical conformation in bacterial membrane-mimetic solutions. It possesses potent broad-spectrum and rapid antimicrobial activity. Meanwhile, it is highly stable and shows low cytotoxicity toward mammalian cells. The microbial killing activity of Hc-CATH is executed through the disruption of cell membrane and lysis of bacterial cells. In addition, Hc-CATH exhibited potent anti-inflammatory activity by inhibiting the LPS-induced production of nitric oxide (NO) and pro-inflammatory cytokines such as TNF-α, IL-1β, and IL-6. Hc-CATH directly binds with LPS to neutralize its toxicity, and it also binds to Toll-like receptor 4 (TLR4/MD2 complex), which therefore inhibits the binding of LPS to TLR4/MD2 complex and the subsequent activation of LPS-induced inflammatory response pathways. Taken together, our study demonstrates that Hc-CATH, the first cathelicidin from sea snake discovered to have both antimicrobial and anti-inflammatory activity, is a potent candidate for the development of peptide antibiotics. 相似文献
77.
Pei Chun Lai Yen Ta Huang Chia Chen Wu Ching-Jung Lai Pen Jung Wang Ted H Chiu 《Journal of biomedical science》2011,18(1):69
Background
Perinatal brain injury is the leading cause of subsequent neurological disability in both term and preterm baby. Glutamate excitotoxicity is one of the major factors involved in perinatal hypoxic-ischemic encephalopathy (HIE). Glutamate transporter GLT1, expressed mainly in mature astrocytes, is the major glutamate transporter in the brain. HIE induced excessive glutamate release which is not reuptaked by immature astrocytes may induce neuronal damage. Compounds, such as ceftriaxone, that enhance the expression of GLT1 may exert neuroprotective effect in HIE. 相似文献78.
多聚ADP核糖聚合酶(PARP)受基因毒剂的特异性诱导。将拟南芥(Arabidopsis thaliana)AtPARP1基因上游长2179bp的启动子片段插入到质粒pAKK687的β-葡萄糖醛酸糖苷酶(GUS)报告基因上游,转化拟南芥。GUS组织化学染色结果表明,GUS报告基因仅在苗龄3-5天的拟南芥根部及花发育早期的雄蕊中表达;1.5μg.mL-1博莱霉素与22μg.mL-1丝裂霉素联用强烈诱导了GUS报告基因的表达(尤其在拟南芥的幼苗和果荚中)。进一步降低抗生素浓度,发现单独使用1μg.mL-1博莱霉素对GUS报告基因也具较强的诱导活性,且对拟南芥幼苗的生长无影响。上述结果表明,AtPARP1启动子是一个新型的具较大应用潜力的抗生素诱导型启动子。 相似文献
79.
Marta De Zotti Barbara Biondi Cristina Peggion Yoonkyung Park Kyung‐Soo Hahm Fernando Formaggio Claudio Toniolo 《Journal of peptide science》2011,17(8):585-594
The medium‐length peptaibiotics are characterized by a primary structure of 14–16 amino acid residues. Despite the interesting antibiotic and antifungal properties exhibited by these membrane‐active peptides, their exact mechanism of action is still unknown. Here, we present our results on heptaibin, a 14‐amino acid peptaibiotic found to exhibit antimicrobial activity against Staphylococcus aureus. We carried out the very challenging synthesis of heptaibin on solid phase and a detailed conformational analysis in solution. The peptaibiotic is folded in a mixed 310‐/α‐helix conformation which exhibits a remarkable amphiphilic character. We also find that it is highly stable toward degradation by proteolytic enzymes and nonhemolytic. Finally, fluorescence leakage experiments using small unilamellar vesicles of three different compositions revealed that heptaibin, although uncharged, is a selective compound for permeabilization of model membranes mimicking the overall negatively charged surface of Gram‐positive bacteria. This latter finding is in agreement with the originally published antimicrobial activity data. Copyright © 2011 European Peptide Society and John Wiley & Sons, Ltd. 相似文献
80.
Pospíšil S Petříčková K Sedmera P Halada P Olšovská J Petříček M 《Journal of applied microbiology》2011,111(5):1116-1128
Aims: Production of minor asukamycin congeners and its new derivatives by combination of targeted genetic manipulations with specific precursor feeding in the producer of asukamycin, Streptomyces nodosus ssp. asukaensis. Methods and Results: Structural variations of manumycins lie only in the diverse initiation of the ‘upper’ polyketide chain. Inactivation of the gene involved in the biosynthesis of cyclohexanecarboxylic acid (CHC) turned off the production of asukamycin in the mutant strain and allowed an increased production of other manumycins with the branched end of the upper chain. The ratio of produced metabolites was further affected by specific precursor feeding. Precursor‐directed biosynthesis of a new asukamycin analogue (asukamycin I, 28%) with linear initiation of the upper chain was achieved by feeding norleucine to the mutant strain. Another asukamycin analogue with the unbranched upper chain (asukamycin H, 14%) was formed by the CHC‐deficient strain expressing a heterologous gene putatively involved in the formation of the n‐butyryl‐CoA starter unit of manumycin A. Conclusions: Combination of the described techniques proved to be an efficient tool for the biosynthesis of minor or novel manumycins. Significance and Impact of the Study: Production of two novel asukamycin derivatives, asukamycins H and I, was achieved. Variations appeared in the upper polyketide chain, the major determinant of enzyme‐inhibitory features of manumycins, affecting their cancerostatic or anti‐inflammatory features. 相似文献