Acta Microbiologica Sinica

[Objective]The aim of this study is to clone and express the nucleotidylytransferase encoding gene-amiE from the biosynthetic gene cluster of amicetin,a disaccharide nucleoside antibiotic,and to characterize AmiE in vitro.[Methods]The amiE,encoding a nucleotidylytransferase of 257 amino acid,was PCR amplified and cloned into pET28a,resulting in the plasmid pCSG4001,which was transformed into E.coli BL21(DE3) for expressing N-(His)6-tag AmiE.The recombinant AmiE was purified by affinity chromatography via AKTA Purifier 10 system.The AmiE-catalyzed reactions were performed using TTP(or UTP) and glucose-1-phosphate as substrates.The enzyme assays were analyzed by HPLC;the substrate flexibility of AmiE was probed with three unnatural sugars-1-phosphate,including galactose-1-phosphate,galactosamine-1-phosphate and mannos-1-phosphate.[Results]The N-(His)6-tag AmiE was expressed in E.coli in soluble form and was successfully purified via Ni+2 mediated affinity chromatography;in vitro biochemical experiments showed that AmiE could convert glucose-1-phosphate into TDP-glucose(or UDP-glucose) in the presence of TTP(or UTP).However,galactose-1-phosphate,galactosamine-1-phosphate and mannos-1-phosphate were not substrates of AmiE.[Conclusion]The amiE was successfully cloned and expressed in E.coli,and the purified AmiE was biochemically confirmed to be a nucleotylyltransferase in amicetin biosynthesis pathway.

Cas Key

ACTA MICROBIOLOGICA SINICA

Cloning,expression and characterization of the nucleotidylyltransferase gene-amiE in amicetin biosynthesis

克隆、表达及友菌素生物合成中核苷酰基转移酶基因amiE的特性分析

[Objective] The characteristics of microscopic distribution and content of Undifilum oxytropis were observed and quantified in different tissues of Oxytropis glabra and Astragalus variabilis from natural grasslands of Inner Mongolia and Ningxia Province. [Methods]Distribution of fungal endophyte was obtained in all the tissues( stems,leaves,seeds and roots) of O. glabra and A. variabilis though paraffin section and staining method of lactic acid phenol cotton blue;and content of fungal endophyte was determined though Real time-qPCR. [Results] Endophytc fungi were observed mainly within the gap between the palisade tissue and parenchymatous tissue of seed coat in seed,mainly colonized in the superficial cells layer near stoma in the leaves,and in pith of stem mainly plants in the parenchymatous tissue around the edge of the vertical axis of the vascular bundle. There was an obviously difference in concentration of U. oxytropis in the plants collected in different locations. The content of U. oxytropis was highest in all seeds of O. glabra,while it was opposite in stems and leaves of two sampling points. Similarly,the content of U. oxytropis was highest in seeds,it was lowest in roots,and stems and leaves were opposite in A. variabilis from two sampling points. The detection limit was0. 029 pg/ng total DNA by Real time-qPCR. [Conclusion] When endophytic fungi infected the tissues of plants,there was selectivity to the tissues and cell type of host,and the colonization and distribution were influenced by habitats in fungal endophytes of locoweeds.

F Universit

Microscopic distribution and quantitative detection of endophytic fungus Undifilum oxytropis from Oxytropis glabra DC and Astragalus variabilis

微观分布及内生真菌Undifilum oxytropis在Oxytropis glabra DC和Astragalus variabilis中的定量检测

[Objective]To investigate the function of glpX gene in mycobacterial physiology and metabolism.[Methods] The glpX mutant of Mycobacterium smegmatis was constructed byusing mycobacteriophage Che9c-encoded recombination system. Then the growth difference between mutant and wild type strain was compared on various carbon sources. Furthermore,we analyzed the glpX gene transcriptional level of wild type strain growing on glucose or oleic acid by realtime PCR. [Results]The growth of M. smegmatis on glycerol or oleic acid is strongly attenuated after the deletion of the glpX gene. The transcription of glpX gene is upregulated in the medium with oleic acid as the sole carbon source. [Conclusion] These results indicate that the fructose 1,6-bisphosphatase encoded by glpX gene is required and nonredundant for mycobacterial gluconeogenesis.

Moh Key

Gene function of glpX in mycobacterium

结核分枝杆菌中glpX基因的功能

[Objective] To clone a halogenase gene from halometabolite-producing Streptomyces sp. 604F to facilitate identification of potential halometabolites and its biosynthetic gene cluster. [Methods] We used agar block method to detect the antimicrobial activity of Streptomyces sp. 604F. We further amplified the conserved regions of type I polyketide synthase( PKS I),type II polyketide synthase( PKS II) and nonribosomal peptide synthetase( NRPS) encoding genes by degenerative PCR. We detected halometabolites in fermentation extracts of Streptomyces sp. 604F analyzed by liquid chromatography-time of flight mass spectrometry( LC-Tof MS). Next,we amplified halogenase gene fragment from Streptomyces sp. 604F by using degenerative primers targeting reduced flavin adenine dinucleotide( FADH2)-dependent halogenase genes. Finally,we cloned the full halogenase gene and its flanking sequences through high-efficiency thermal asymmetric interlaced PCR( hiTAIL-PCR). [Results]Streptomyces sp. 604F showed promising antifungal activity against Candida albicans ATCC 10231,and its genome contained genes encoding PKS I,PKS II,NRPS and a halogenase with1443 bp. The halogenase is a new non-tryptophan halogenase,and most closely related to halogenases catalyzing the chlorination of glycopeptides. [Conclusion] Streptomyces sp. 604F possessed a new non-tryptophan halogenase,which may be involved in halogenation of glycopeptide-like metabolites. The cloning and analysis of this halogenase have provided guidance for searching target halometabolites,and laid the foundation for obtaining the biosynthetic gene cluster.

Soa Key

Cloning and analysis of a halogenase gene of Streptomyces sp. 604F from the Arctic Ocean

北极洋来源链霉菌604F卤化酶基因的克隆与分析

Xiao XiaoPeng

Jin Peng

Cai MinMin

Zheng LongYu

Li Wu

Yu ZiNiu

Zhang JiBin

Chicken manure co-conversion by Hermetia illucens larvae with non-associated microorganism.

黑水虻幼虫协同非关联微生物转化鸡粪。

[Objective] Twenty-nine healthy young volunteers from Hainan were selected as the research objects to analyze the diversity of gut microbes and to explore the effects of Lactobacillus paracasei subsp. paracasei Zhang on the fecal microbiota. [Methods] Volunteers were supplemented with 1×1010 CFU LCZ powders per day for 14 days. Fecal samples were collected on 0, 14 and 28 days. Microbial composition and structure of fecal were analyzed by PacBio SMRT. [Results] Firmicutes and Bacteroidetes were the most abundant phylum. Meanwhile, Bacteroides were the predominant genus. At the species level, the dominant bacteria species were Faecalibacterium prausnitzii, Eubacterium rectale and Prevotella copri. After taking LCZ for 14 days, the dominant bacteria did not change significantly, while low-abundance bacteria such as Oscillospira and Parabacteroides were significantly increased（P<0.05）. In addition, volunteers were divided into two groups according to the changes of Lactobacillus. Butyricicoccus pullicaecorum, Lactococcus raffinolactis and Parabacteroides distasoni were significantly increased in samples of those with significantly increased content of Lactobacillus. Inversely, volunteers with no significant changes in the Lactobacillus content in their gut did not find similar changes. [Conclusion] Continuous consumption of probiotics LCZ for 2 weeks had no significant effect on the relative abundance of dominant bacteria in fecal microbiota of volunteers, while the low-abundance bacteria change significantly.

Lingling Shen

Haiyan Xu

Qiangchuan Hou

Feiyan Zhao

Yahua Liu

Jiao Wang

Zhihong Sun

Heping Zhang

Effect of probiotics Lactobacillus paracasei subsp. paracasei Zhang on gut microbiota composition of young people living in Hainan

益生菌副干酪乳杆菌副干酪乳杆菌张氏对海南年轻人肠道菌群组成的影响

益生菌副干酪乳杆菌副干酪亚种张氏对海南年轻人肠道菌群组成的影响

益生菌副干酪乳杆菌张氏亚种对海南年轻人肠道菌群组成的影响

益生菌副干酪乳杆菌副干酪亚种张氏菌对海南年轻人肠道菌群组成的影响

益生菌副干酪乳杆菌副干酪亚种张氏菌对海南青年肠道菌群组成的影响

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Cloning,expression and characterization of the nucleotidylyltransferase gene-amiE in amicetin biosynthesis

Microscopic distribution and quantitative detection of endophytic fungus Undifilum oxytropis from Oxytropis glabra DC and Astragalus variabilis

Gene function of glpX in mycobacterium

Cloning and analysis of a halogenase gene of Streptomyces sp. 604F from the Arctic Ocean

Chicken manure co-conversion by Hermetia illucens larvae with non-associated microorganism.

Effect of probiotics Lactobacillus paracasei subsp. paracasei Zhang on gut microbiota composition of young people living in Hainan

Effect of probiotics Lactobacillus paracasei subsp. paracasei Zhang on gut microbiota composition of young people living in Hainan

Effect of probiotics Lactobacillus paracasei subsp. paracasei Zhang on gut microbiota composition of young people living in Hainan

Effect of probiotics Lactobacillus paracasei subsp. paracasei Zhang on gut microbiota composition of young people living in Hainan

Effect of probiotics Lactobacillus paracasei subsp. paracasei Zhang on gut microbiota composition of young people living in Hainan

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