Antibacterial and antitubercular activity of fosmidomycin, FR900098, and their lipophilic analogs

The nonmevalonate pathway (NMP) of isoprene biosynthesis is an exciting new route toward novel antibiotic development. Inhibitors against several enzymes in this pathway are currently under examination. A significant liability of many of these agents is poor cell penetration. To overcome and improve our understanding of this problem, we have synthesized a series of lipophilic, prodrug analogs of fosmidomycin and FR900098, inhibitors of the NMP enzyme Dxr. Several of these compounds show improved antibacterial activity against a panel of organisms relative to the parent compound, including activity against Mycobacterium tuberculosis (Mtb). Our results show that this strategy can be an effective way for improving whole cell activity of NMP inhibitors.     

Evidence of artemisinin production from IPP stemming from both the mevalonate and the nonmevalonate pathways

The potent antimalarial sesquiterpene lactone, artemisinin, is produced in low quantities by the plant Artemisia annua L. The source and regulation of the isopentenyl diphosphate (IPP) used in the biosynthesis of artemisinin has not been completely characterized. Terpenoid biosynthesis occurs in plants via two IPP-generating pathways: the mevalonate pathway in the cytosol, and the non-mevalonate pathway in plastids. Using inhibitors specific to each pathway, it is possible to resolve which supplies the IPP precursor to the end product. Here, we show the effects of inhibition on the two pathways leading to IPP for artemisinin production in plants. We grew young (7–14 days post cotyledon) plants in liquid culture, and added mevinolin to the medium to inhibit the mevalonate pathway, or fosmidomycin to inhibit the non-mevalonate pathway. Artemisinin levels were measured after 7–14 days incubation, and production was significantly reduced by each inhibitor compared to controls, thus, it appears that IPP from both pathways is used in artemisinin production. Also when grown in miconazole, an inhibitor of sterol biosynthesis, there was a significant increase in artemisinin compared to controls suggesting that carbon was shifted from sterols into sesquiterpenes. Collectively these results indicate that artemisinin is probably biosynthesized from IPP pools from both the plastid and the cytosol, and that carbon from competing pathways can be channeled toward sesquiterpenes. This information will help advance our understanding of the regulation of in planta production of artemisinin.     

炭样小单孢菌中抗生素生物合成相关蛋白的筛选

目的：筛选一株具有广谱抗菌活性的炭样小单孢菌JXNU-1中核苷类抗生素生物合成相关蛋白。方法：通过iTRAQ定量蛋白质组学技术对JXNU-1菌体生长期（36h）和产物合成期（108h）的差异蛋白进行鉴定和功能分析。结果：基于iTRAQ定量蛋白质组学技术共鉴定出炭样小单孢菌总蛋白质2390个,差异表达蛋白172个,在产物合成期（108h）表达上调76个、表达下调96个。通过蛋白GO和COG注释等功能分析,筛选出12个与抗生素合成密切相关蛋白和5个生物合成基因簇。结论：利用iTRAQ技术筛选出炭样小单孢菌JXNU-1的抗生素合成相关蛋白,为阐明该抗生素的生物合成机制奠定实验依据。     

The high resolution structure of tyrocidine A reveals an amphipathic dimer

Tyrocidine A, one of the first antibiotics ever to be discovered, is a cyclic decapeptide that binds to membranes of target bacteria, disrupting their integrity. It is active against a broad spectrum of Gram-positive organisms, and has recently engendered interest as a potential scaffold for the development of new drugs to combat antibiotic-resistant pathogens. We present here the X-ray crystal structure of tyrocidine A at a resolution of 0.95 Å. The structure reveals that tyrocidine forms an intimate and highly amphipathic homodimer made up of four beta strands that associate into a single, highly curved antiparallel beta sheet. We used surface plasmon resonance and potassium efflux assays to demonstrate that tyrocidine binds tightly to mimetics of bacterial membranes with an apparent dissociation constant (K_D) of 10 μM, and efficiently permeabilizes bacterial cells at concentrations equal to and below the K_D. Using variant forms of tyrocidine in which the fluorescent probe p-cyano-phenylalanine had been inserted on either the polar or apolar face of the molecule, we performed fluorescence quenching experiments, using both water-soluble and membrane-embedded quenchers. The quenching results, together with the structure, strongly support a membrane association model in which the convex, apolar face of tyrocidine's beta sheet is oriented toward the membrane interior, while the concave, polar face is presented to the aqueous phase.     

Molecular analysis of tlrD, an MLS resistance determinant from the tylosin producer, Streptomyces fradiae

The macrolide antibiotic, tylosin (Ty), is produced by Streptomyces fradiae. Two resistance determinants (tlrA, synonym ermSF, and tlrD) conferring resistance to macrolide, lincosamide and streptogramin B type (MLS) antibiotics were previously isolated from this strain, and their products shown to methylate 23S ribosomal RNA (rRNA) at a common site, thereby rendering the ribosomes MLS resistant. However, the T1rA and T1rD proteins differ in their action; the former dimethylates, and the latter monomethylates, the target nucleotide. Here, 2.2 kb of DNA from the tylLM region of the tylosin biosynthetic gene cluster of S. fradiae has been sequenced and shown to encompass tlrD. Comparison of the sequences of tlrA and tlrD (and of their deduced products) with those of related (‘erm-type’) genes from other actinomycetes suggests that the combined presence of tlrA and tlrD in S. fradiae is not the result of recent gene duplication.     

幽门螺杆菌抗生素耐药机制研究进展

幽门螺杆菌(Helicobacter pylori,H.pylori)感染可引起消化性溃疡、胃粘膜相关淋巴组织淋巴瘤和胃癌。随着抗生素耐药性的问题越来越严重,耐药机制的研究也不断深入。分子检测方法,尤其是核酸检测技术,可高效、快速、准确地检测幽门螺杆菌抗生素耐药基因及突变,对幽门螺杆菌感染的临床治疗发挥重要的指导作用,同时也可对幽门螺杆菌抗生素耐药性进行大规模及时有效监控。本文讨论了关于幽门螺杆菌抗生素耐药机制并着重总结了相关耐药基因及突变。     

Phosphorylation of chloramphenicol by a recombinant protein Yhr2 from Streptomyces avermitilis MA4680

Although phosphorylation of chloramphenicol has been shown to occur in the chloramphenicol producer, Streptomyces venezuelae, there are no reports on the existence of chloramphenicol phosphorylase in other Streptomyces species. In the present study, we report the modification of chloramphenicol by a recombinant protein, designated as Yhr2 (encoded by SAV_877), from Streptomyces avermitilis MA4680. Recombinant Yhr2 was expressed in Escherichia coli BL21 (DE3) and the cells expressing this recombinant protein were shown to phosphorylate chloramphenicol to a 3′-O-phosphoryl ester derivative, resulting in an inactivated form of the antibiotic. Expression of yhr2 conferred chloramphenicol resistance to E. coli cells up to 25 μg/mL and in an in vitro reaction, adenosine triphosphate (ATP), guanosine triphosphate (GTP), adenosine diphosphate (ADP) and guanosine diphosphate (GDP) were shown to be the phosphate donors for phosphorylation of chloramphenicol. This study highlights that antibiotic resistance conferring genes could be easily expressed and functionalized in other organisms that do not produce the respective antibiotic.     

抗生素产生菌诱变育种的研究进展

一种抗生素能否产业化决定于抗生素产生菌菌种的产素水平。因而如何提高抗生素产生菌的产素水平成为抗生素研究工作者长期不懈探索的重要课题。其中,诱变育种以其技术设备简单、省时省力等优点得到了微生物育种工作者的青睐,在诱变育种技术上取得了巨大进展,为抗生素工业化生产发挥了重要作用。综述了抗生素产生菌的诱变育种研究进展。     

Factors affecting antimicrobial activity of Synechococcus leopoliensis

An antimicrobial agent is produced by the cyanobacterium Synechococcus leopoliensis which was found to be active against the Gram-positive bacterium Staphylococcus aureus. The effects of temperature, pH, incubation period, some media and different nitrogen and carbon sources on both growth and antimicrobial activity were investigated. Temperature 35 °C and pH 8 were the best for growth and antimicrobial agent production and 14 and 15 days of incubation were found to be the best for maximum growth and antimicrobial activity, respectively, in the medium BG-11.

No antimicrobial activity could be detected by the use of G medium, moderate activity was recorded with Chu 10 medium, while high activity was reported in BG-11 medium. Leucine was the best nitrogen source for antimicrobial activity, while maximum antimicrobial activity was introduced by using the carbon sources, citrate and acetate. Very high antimicrobial activity could be detected by using the carbon source galactose in combination with the nitrogen source alanine or by using arabinose with methionine.     

A 15N nuclear magnetic resonance study of the biosynthesis of quinoxaline antibiotics

Uniformly ¹⁵N-labelled triostin A and echinomycin have been prepared by growing the producing organisms on enriched media and their ¹⁵N nuclear magnetic resonance spectra partially assigned by a combination of nuclear Overhauser effect and scalar coupling constant measurements. Selective feeding experiments using unlabelled L-tryptophan-supplemented media have shown that N-1 and N-4 of the quinoxaline rings have their origins in the indole and amino groups of tryptophan, respectively.     

球形芽孢杆菌(Bacillus sphaericus)对抗生素的抗性

考查了27株球形芽孢杆菌有毒株对青霉素、链霉素、四环素、氯霉素、红霉素、卡那霉素、庆大霉素、麦迪霉素、核糖霉素、利福平、吖啶黄、磺胺和迭氮钠的自然抗性。结果表明,所有菌株对高浓度的链霉素和迭氮钠及低浓度的氯霉素有抗性,大部分菌株对四环素有抗性。所有菌株对所考查的其它几种抗生素敏感。     

口腔血链球菌质粒的初步研究

口腔血链球菌是牙周主要有益菌,质粒是细胞染色体外辅助性遗传物质可以赋予细胞不同表型。本研究目的提检测血链球菌质粒ＤＮＡ存在情况,鉴定其分子大小,初步了解血链菌质粒与耐药性的关系,为今后更深入地在分子水平研究血链菌奠定实验基础。     

山苍子油对霉菌抗菌性及其与黄曲霉产毒关系的研究

采用平板法比较天然增香剂山苍子油与合成食用防腐剂苯甲酸钠、山梨酸钾对8种霉菌的抗菌效力。结果表明,在培养基pH4．5时山苍子油对多数霉菌的最低抑菌浓度为1．77mg／ml,与山梨酸钾的抑菌强度相近,比苯甲酸钠强;但当培养基pH5．5以上时苯甲酸钠对霉菌几乎无效,山梨酸钾的抗菌效力也有减弱,而山苍子油受影响很小,其活性pH范围为4．5～8．5。山苍子油用脂肪酸蔗糖酯乳化,其抗菌效力增强一倍。同时,从山苍子油与黄曲霉产毒关系的实验中发现,山苍子油对黄曲霉产生黄曲霉毒素有较强的抑制作用。     

利用胞内代谢物变化预测大肠埃希菌耐药性

目的利用胞内代谢物量的差异达到预测大肠埃希氏菌的药物敏感性结果。方法收集临床分离大肠埃希菌120株,在头孢他啶存在条件下培养4 h。收集菌体,测定细胞内化合物然后进行多变量分析。结果 120株大肠埃希菌的聚类分析结果很好地体现了药物敏感性特征,57株耐药菌中,有5株被误判,准确率为91%;47株中敏菌株中,有8株被误判,准确率为83%;16株敏感菌中有2株被误判;准确率为88%。结论该方法能够有效预测菌株的药敏结果,而且快速可靠。