Journal of Microbiology

Journal Article

Fresh Washed Microbiota Transplantation Alters Gut Microbiota Metabolites to Ameliorate Sleeping Disorder Symptom of Autistic Children: Nai-Hua Liu , Hong-Qian Liu , Jia-Yi Zheng , Meng-Lu Zhu , Li-Hao Wu , Hua-Feng Pan , Xing-Xiang He; J. Microbiol. 2023;61(8):741-753. Published online September 4, 2023; DOI: https://doi.org/10.1007/s12275-023-00069-x

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Accumulating studies have raised concerns about gut dysbiosis associating autism spectrum disorder (ASD) and its related symptoms. However, the effect of gut microbiota modification on the Chinese ASD population and its underlying mechanism were still elusive. Herein, we enrolled 24 ASD children to perform the first course of fresh washed microbiota transplantation (WMT), 18 patients decided to participate the second course, 13 of which stayed to participate the third course, and there were 8 patients at the fourth course. Then we evaluated the effects of fresh WMT on these patients and their related symptoms. Our results found that the sleeping disorder symptom was positively interrelated to ASD, fresh WMT significantly alleviated ASD and its sleeping disorder and constipation symptoms. In addition, WMT stably and continuously downregulated Bacteroides/ Flavonifractor/Parasutterella while upregulated Prevotella_9 to decrease toxic metabolic production and improve detoxification by regulating glycolysis/myo-inositol/D-glucuronide/D-glucarate degradation, L-1,2-propanediol degradation, fatty acid β-oxidation. Thus, our results suggested that fresh WMT moderated gut microbiome to improve the behavioral and sleeping disorder symptoms of ASD via decrease toxic metabolic production and improve detoxification. Which thus provides a promising gut ecological strategy for ASD children and its related symptoms treatments.

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Untargeted urine metabolomics and machine learning provide potential metabolic signatures in children with autism spectrum disorder
Xian Liu, Xin Sun, Cheng Guo, Zhi-Fang Huang, Yi-Ru Chen, Fang-Mei Feng, Li-Jie Wu, Wen-Xiong Chen
Frontiers in Psychiatry.2024;[Epub] CrossRef
Washed Microbiota Transplantation Improves the Sleep Quality in Patients with Inflammatory Bowel Disease
Qianqian Li, Yujie Liu, Zulun Zhang, Sheng Zhang, Xiao Ding, Faming Zhang
Nature and Science of Sleep.2024; Volume 16: 1141. CrossRef

Research Support, Non-U.S. Gov'ts

Expression, Purification, and Characterization of Recombinant Fibulin-5 in a Prokaryote Expression System: Myoung Seok Jeong , Chang Soo Kang , Yeon Soo Han , In Seok Bang; J. Microbiol. 2010;48(5):695-700. Published online November 3, 2010; DOI: https://doi.org/10.1007/s12275-010-0320-6

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Abstract

Fibulin-5 is a widely expressed, integrin-binding extracellular matrix protein that mediates endothelial cell adhesion and scaffolds cells to elastic fibers. To investigate anti-angiogenesis activities and context-specific activities on responsive cells of recombinant fibulin-5 (rfibulin-5) expressed in Escherichia coli, the cDNA of fibulin-5 cloned from a human placenta cDNA library was inserted into the pET32a (+) vector to allow fibulin-5 expression as a Trx fusion protein. The fusion protein Trx-fibulin-5, expressed as insoluble inclusion bodies, was solubilized and its resulting expression level reached to 15% of the total cell protein. The Trxfibulin-5 was purified effectively by N2+-chelating chromatography and then identified by Western blotting analysis with an anti-His tag antibody. The purified Trx-fibulin-5 was refolded by dialysis against redox reagents, and the rfibulin-5 released from the fusion protein by enterokinase cleavage was purified using a RESOURCE RPC column. The final purified rfibulin-5 effectively inhibited angiogenesis in chicken embryos in a dose-dependent manner through a chorioallantoic membrane (CAM) assay. Additionally, rfibulin-5 potently suppressed in vitro proliferation of human umbilical vein endothelial cells, but stimulated that of human dermal fibroblasts. The expression and in vitro refolding of rfibulin-5 resulted in production of an active molecule with a yield of 2.1 mg/L.

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Impact of UV pre-treatment on the Longissimus thoracis et lumborum muscle proteomes of dry-aged beef cuts: A characterisation within two sampling locations
Sara Álvarez, Carlos Álvarez, Anne Maria Mullen, Eileen O'Neill, Mohammed Gagaoua
Meat Science.2025; 221: 109729. CrossRef
Genome-wide association study on reproductive traits in Jinghai Yellow Chicken
G.X. Zhang, Q.C. Fan, J.Y. Wang, T. Zhang, Q. Xue, H.Q. Shi
Animal Reproduction Science.2015; 163: 30. CrossRef
Production and on-column re-folding of human vascular endothelial growth factor 165 in Escherichia coli
Sun Kwon Bang, Young Sik Kim, Byung Soo Chang, Cheol Beom Park, In Seok Bang
Biotechnology and Bioprocess Engineering.2013; 18(5): 835. CrossRef

Fusion Expression and Immunogenicity of EHEC EspA-Stx2A1 Protein: Implications for the Vaccine Development: Yan Cheng , Youjun Feng , Ping Luo , Jiang Gu , Shu Yu , Wei-jun Zhang , Yan-qing Liu , Qing-xu Wang , Quan-ming Zou , Xu-hu Mao; J. Microbiol. 2009;47(4):498-505. Published online September 9, 2009; DOI: https://doi.org/10.1007/s12275-009-0116-8

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Abstract: Shiga toxin 2 (Stx2) is a major virulence factor for enterohemorrhagic Escherichia coli (EHEC), which is encoded by λ lysogenic phage integrated into EHEC chromosome. Stx2A1, A1 subunit of Stx2 toxin has gathered extensive concerns due to its potential of being developed into a vaccine candidate. However, the substantial progress is hampered in part for the lack of a suitable in vitro expression system. Here we report use of the prokaryotic system pET-28a::espA-Stx2A1/BL21 to carry out the fusion expression of Stx2A1 which is linked to E. coli secreted protein A (EspA) at its N-terminus. Under the IPTG induction, EspA- Stx2A1 fusion protein in the form of inclusion body was obtained successfully, whose expression level can reach about 40% of total bacterial protein at 25°C, much higher than that at 37°C. Western blot test suggested the refolded fusion protein is of excellent immuno-reactivity with both monoclonal antibodies, which are specific to EspA and Stx2A1, respectively. Anti-sera from Balb/c mice immunized with the EspA-Stx2A1 fusion protein were found to exhibit strong neutralization activity and protection capability in vitro and in vivo. These data have provided a novel feasible method to produce Stx2A1 in large scale in vitro, which is implicated for the development of multivalent subunit vaccines candidate against EHEC O157:H7 infections.

Stable Expression and Secretion of Polyhydroxybutyrate Depolymerase of Paucimonas lemoignei in Escherichia coli: Se Whan Park , Moon Gyu Chung , Hwa Young Lee , Jeong Yoon Kim , Young Ha Rhee; J. Microbiol. 2008;46(6):662-669. Published online December 24, 2008; DOI: https://doi.org/10.1007/s12275-008-0283-z

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Abstract: An efficient strategy for the expression and secretion of extracellular polyhydroxybutyrate depolymerase (PhaZ1) of Paucimonas lemoignei in Escherichia coli was developed by employing the signal peptide of PhaZ1 and a truncated ice nucleation protein anchoring motif (INPNC). Directly synthesized mature form of PhaZ1 was present in the cytoplasm of host cells as inclusion bodies, while a construct containing PhaZ1 and its own N-terminal signal peptide (PrePhaZ1) enabled the secretion of active PhaZ1 into the extracellular medium. However, the PrePhaZ1 construct was harmful to the host cell and resulted in atypical growth and instability of the plasmid during the cultivation. In contrast, INPNC-PhaZ1 and INPNCPrePhaZ1 fusion constructs did not affect growth of host cells. INPNC-PhaZ1 was successfully displayed on the cell surface with its fusion form, but did not retain PhaZ1 activity. In the case of INPNC-PrePhaZ1, the initially synthesized fusion form was separated by precise cleavage of the signal peptide, and active PhaZ1 was consequently released into the culture medium. The amount of PhaZ1 derived from E. coli (INPNC-PrePhaZ1) was almost twice as great as that directly expressed from E. coli (PrePhaZ1), and was predominantly (approximately 85%) located in the periplasm when cultivated at 22°C but was efficiently secreted into the extracellular medium when cultivated at 37°C.

Expression of a Recombinant Cry1Ac Crystal Protein Fused with a Green Fluorescent Protein in Bacillus thuringiensis subsp. kurstaki Cry-B: Jong Yul Roh , In Hee Lee , Ming Shun Li , Jin Hee Chang , Jae Young Choi , Kyung Saeng Boo , Yeon Ho Je; J. Microbiol. 2004;42(4):340-345.; DOI: https://doi.org/2101 [pii]

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Abstract: To investigate the co-expression and crystallization of a fusion gene between the Bacillus thuringiensis crystal protein and a foreign protein in B. thuringiensis, the expression of the Cry1Ac fused with green fluorescent protein (GFP) genes in a B. thuringiensis Cry-B strain was examined. The cry1Ac gene was cloned in the B. thuringiensis-E. coli shuttle vector, pHT3101, under the control of the native cry1Ac gene promoter, while the GFP gene was inserted into the XhoI site upstream of the proteolytic cleavage site, in the middle region of the cry1Ac gene (pProAc-GFP). The B. thuringiensis Cry-B strain carrying pProAc-GFP (ProAc-GFP/CB) did not produce any inclusion bodies. However, the transformed strain expressed fusion protein forms although the expression level was relatively low. Furthermore, an immunoblot analysis using GFP and Cry1Ac antibodies showed that the fusion protein was not a single species, but rather multiple forms. In addition, the N-terminal fragment of Cry1Ac and a non-fused GFP were also found in the B. thuringiensis Cry-B strain after autolysis. The sporulated cells before autolysis and the spore-crystal mixture after autolysis of ProAc-GFP/CB exhibited insecticidal activities against Plutella xylostella larvae. Accordingly, the current results suggest that a fusion crystal protein produced by the transfomant, ProAc-GFP/CB, can be functionally expressed but easily degraded in B. thuringiensis.

Identification of a cellular protein interacting with murine retrovirus Gag polyproteins: Choi , Won Ja; J. Microbiol. 1996;34(4):311-315.

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Abstract: The retroviral Gag polyprotein directs the assembly of virion particles and plays an important role in some events after entry into a host cell. The Gag polyprotein of a virus mixture is responsible for inducing murine acquired immunodeficiency syndrome (MAIDS) when injected into susceptible strains of mice. In order to identify the host cellular proteins which interact with the MAIDS virus Gag proteins and possibly mediate the function of the Gag proteins, mouse T-cell leukemic cDNA expression library was screened using the yeast GAL4 two hybrid system. Of 11 individual positive clones, the clone Y1 was selected for the study of protein-protein interaction. Its DNA sequence revealed that it was an exact match to the murine SH3 domain-containing protein SH3P8. It is expressed as 2.4 kbp transcripts in testis at higher levels and in various tissues tested at lower levels. Glutathione S-transferase-Y1 fusion protein binds tightly to Pr60^def-gag as well as Pr 65^eco-gag.

Production of the Nucleocapsid Protein of Newcastle Disease Virus in Escherichia coli and its Assembly into Ring- and Nucleocapsid-like Particles: Chiew Ling Kho , Wen Siang Tan , Khatijah Yusoff; J. Microbiol. 2001;39(4):293-299.

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Abstract: The nucleocapsid (NP) protein of Newcastle disease virus (NDV) and its derivative (NP_cfus ) containing the myc region and six histidine residues fused to its C-terminus were expressed abundantly in Escherichia coli. The proteins were purified by sucrose gradient centrifugation. Both the NP and NP_cfus proteins self-assembled into ring-like particles with a diameter of 24 +- 2 nm around a central hole of 7 +- 1 nm. Some of these ring-like particles stacked together to form nucleocapsid-like structures which are heterogeneous in length with a diameter of 20 +- 2 nm and a central hollow of 5 +- 1 nm. Only a very small amount of the monomers in the particles was linked by inter-molecular disulfide bonds. Fusion of the C-terminal end to 29 amino acids inclusive of the myc epitope and His-tag did not impair ring assembly but inhibited the formation of the long herringbone structures. Immunogold labeling of the particles with the anti-myc antibody showed that the C-terminus of the NP_cfus protein is exposed on the surface of these ring-like particles.

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