MICROBIAL PATHOGENESIS
·
2021
·
IF 3.738
Xiufen Du,Daquan Tan,Yang Gong,Yifan Zhang,Jiangyuan Han,Wei Lv,Tao Xie,Pu He,Zongjie Hou,Kun Xu,Jiying Tan,Bingdong Zhu
Protein-based subunit vaccine against tuberculosis (TB) is regarded as safer but with lower immunogenicity . To investigate effective adjuvant to improve the immunogenicity of TB subunit vaccine, we modified ploy(I:C) onto PLGA-PEG copolymer nanoparticle with polydopamine to produce a new nanoparticle adjuvant named “PLGA-PEG-poly(I:C)” (NP). M. tuberculosis fusion proteins Mtb10.4-HspX and ESAT-6-Rv2626c (M4) were encapsulated in the nanoparticles to produce the NP/M4 subunit vaccine. The PLGA-PEG/M4 nanoparticle was 200.21 ± 1.07 nm in diameter, and the polydispersity index (PDI) was 0.127 ± 0.02. Following modification with poly(I:C) by polydopamine, the NP/M4 was administered to C57BL/6 female mice intranasally and the immune responses were evaluated. The NP/M4 significantly induced antigen-specific CD4 + T cells proliferation, IL-2 and IFN-γ production. In addition, the NP/M4 could promote the production of antigen-specific IgG, IgG1 , IgG2c in serum, and sIgA in lung washings . Overall, our results indicated that the NP would be a potential TB subunit vaccine adjuvant with the ability to induce strong Th1-type cell-mediated immunity and humoral immune responses .
研究领域:
免疫学
·
疫苗学
·
纳米技术
·
结核病
✓ 已验证
DOI
PubMed
高影响因子
Frontiers in Cellular and Infection Microbiology
·
2019
·
IF 3.518
Chen Changming,Han Xiuyan,Yan Qiulong,Wang Chao,Jia Liqiu,Taj Ayaz,Zhao Lizhe,Ma Yufang
Mycobacterium tuberculosis bifunctional enzyme GlmU is a novel target for anti-TB drugs and is involved in glycosyl donor UDP-N-acetylglucosamine biosynthesis. Here, we found that TPSA (2-[5-(2-{[4-(2-thienyl)-2-pyrimidinyl]sulfanyl}acetyl)-2-thienyl]acetic acid) was a novel inhibitor for GlmU acetyltransferase activity (IC50: 5.3 μM). The interaction sites of GlmU and TPSA by molecular docking were confirmed by site-directed mutagenesis. TPSA showed an inhibitory effect on Mtb H37Ra growth and intracellular H37Ra in macrophage cells (MIC: 66.5 μM). To investigate why TPSA at a higher concentration (66.5 μM) was able to inhibit H37Ra growth, proteome and transcriptome of H37Ra treated with TPSA were analyzed. The expression of two methyltransferases MRA_0565 (Rv0558) and MRA_0567 (Rv0560c) were markedly increased. TPSA was pre-incubated with purified Rv0558 and Rv0560c in the presence of S-adenosylmethionine (methyl donor) respectively, resulting in its decreased inhibitory effect of GlmU on acetyltransferase activity. The inhibition of TPSA on growth of H37Ra with overexpressed Rv0558 and Rv0560c was reduced. These implied that methyltransferases could modify TPSA. The methylation of TPSA catalyzed by Rv0560c was subsequently confirmed by LC-MS. Therefore, TPSA as a GlmU acetyltransferase activity inhibitor may offer a structural basis for new anti-tuberculosis drugs. TPSA needs to be modified further by some groups to prevent its methylation by methyltransferases.
研究领域:
微生物学
·
药理学
·
分子生物学
✓ 已验证
DOI
PubMed
高影响因子
PROCESS BIOCHEMISTRY
·
2026
·
IF 4.4
Liyang Xu,Yujia Wang,Feiyu Yan,Wenjing Pan,Yingjia Tong,Jinsong Gong,Xiaomei Zhang,Jinsong Shi,Zhenghong Xu,Hui Li
Mycolicibacterium is a crucial industrial host for steroid biotransformation, yet its genetic engineering has long been limited by inefficient homologous recombination (HR) and strong non-homologous end joining (NHEJ) activity. Here, we established and optimized a suicide plasmid-based genome editing system for Mycobacterium sp. LY-1. The constructed vector p2NILB, combining kanamycin selection with sacB-mediated counter-selection, enabled precise and markerless allelic exchange. Suppression of NHEJ through deletion of ku or ligD , together with optimization of homology arm length, increased gene knockout efficiency to over 70%. This optimized platform was subsequently applied to metabolic engineering of sterol degradation to enhance 9α-hydroxyandrostenedione (9α-OH-AD) biosynthesis. Sequential deletion of five kstD isoenzymes blocked downstream steroid degradation, with kstD1 identified as the major contributor to product loss. Functional analysis revealed KshA2 as the key 9α-hydroxylase, and its overexpression increased the 9α-OH-AD titer to 3.41 g/L. In addition, deletion of the branched-chain acid transporter gene mmpL3 further enhanced 9α-OH-AD production to 4.19 g/L. To overcome substrate utilization limitations under high phytosterol loading conditions, a soybean oil‑Tween 80 solvent system overcame substrate limitation, enabling 15.71 g/L 9α‑OH‑AD from 30 g/L phytosterols. This study provides a robust genome editing framework and an effective strategy for engineering steroid-producing actinobacteria.
✓ 已验证
DOI
高影响因子
Microbial Cell Factories
·
2020
·Vol. 19
·期 1
·1-11
·
IF 4.187
Xiong Liang-Bin,Liu Hao-Hao,Zhao Ming,Liu Yong-Jun,Song Lu,Xie Zhi-Yong,Xu Yi-Xin,Wang Feng-Qing,Wei Dong-Zhi
Background The bioconversion of phytosterols into high value-added steroidal intermediates, including the 9 α -hydroxy-4-androstene-3,17-dione (9-OHAD) and 22-hydroxy-23,24-bisnorchol-4-ene-3-one (4-HBC), is the cornerstone in steroid pharmaceutical industry. However, the low transportation efficiency of hydrophobic substrates into mycobacterial cells severely limits the transformation. In this study, a robust and stable modification of the cell wall in M. neoaurum strain strikingly enhanced the cell permeability for the high production of steroids. Results The deletion of the nonessential kasB , encoding a β-ketoacyl-acyl carrier protein synthase, led to a disturbed proportion of mycolic acids (MAs), which is one of the most important components in the cell wall of Mycobacterium neoaurum ATCC 25795. The determination of cell permeability displayed about two times improvement in the kasB -deficient strain than that of the wild type M. neoaurum . Thus, the deficiency of kasB in the 9-OHAD-producing strain resulted in a significant increase of 137.7% in the yield of 9 α -hydroxy-4-androstene-3,17-dione (9-OHAD). Ultimately, the 9-OHAD productivity in an industrial used resting cell system was reached 0.1135 g/L/h (10.9 g/L 9-OHAD from 20 g/L phytosterol) and the conversion time was shortened by 33%. In addition, a similar self-enhancement effect (34.5%) was realized in the 22-hydroxy-23,24-bisnorchol-4-ene-3-one (4-HBC) producing strain. Conclusions The modification of kasB resulted in a meaningful change in the cell wall mycolic acids. Deletion of the kasB gene remarkably improved the cell permeability, leading to a self-enhancement of the steroidal intermediate conversion. The results showed a high efficiency and feasibility of this construction strategy.
研究领域:
生物化学
·
微生物学
·
制药工程
✓ 已验证
产品关联
DOI
PubMed
高影响因子
APOPTOSIS
·
2016
·Vol. 22
·期 4
·502-509
·
IF 3.592
Yang Wenmin,Deng Wanyan,Zeng Jie,Ren Sai,Ali Md Kaisar,Gu Yinzhong,Li Yangyuling,Xie Jianping
Mycobacterium tuberculosis PE/PPE family proteins, named after the presence of conserved PE (Pro-Glu) and PPE (Pro-Pro-Glu) domains at N-terminal, are prevalent in M. tuberculosis genome. The function of most PE/PPE family proteins remains elusive. To characterize the function of PE_PGRS18, the encoding gene was heterologously expressed in M. smegmatis , a nonpathogenic mycobacterium. The recombinant PE_PGRS18 is cell wall associated. M. smegmatis PE_PGRS18 recombinant showed differential response to stresses and altered the production of host cytokines IL-6, IL-1β, IL-12p40 and IL-10, as well as enhanced survival within macrophages largely via attenuating the apoptosis of macrophages. In summary, the study firstly unveiled the role of PE_PGRS18 in physiology and pathogenesis of mycobacterium.
研究领域:
微生物学
·
免疫学
·
分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Scientific Reports
·
2017
·
IF 4.259
Wanyan Deng,Quanxin Long,Jie Zeng,Ping Li,Wenmin Yang,Xinchun Chen,Jianping Xie
The success of Mycobacterium tuberculosis (M. tuberculosis) as a pathogen is largely contributes to its ability to manipulate the host immune responses. The genome of M. tuberculosis encodes multiple immune-modulatory proteins, including several members of the multi-genic PE_PPE family. Despite of intense research, the roles of PE_PGRS proteins in mycobacterial pathogenesis remain elusive. The function of M. tuberculosis PE_PGRS41, characterized by an extended and unique C-terminal domain, was studied. Expression of PE_PGRS41 in Mycobacterium smegmatis, a non-pathogenic species intrinsically deficient of PE_PGRS, severely impaired the resistance of the recombinant to multiple stresses via altering the cell wall integrity. Macrophages infected by M. smegmatis harboring PE_PGRS41 decreased the production of TNF-α, IL-1β and IL-6. In addition, PE_PGRS41 boosted the survival of M. smegmatis within macrophage accompanied with enhanced cytotoxic cell death through inhibiting the cell apoptosis and autophagy. Taken together, these results implicate that PE_PGRS41 is a virulence factor of M. tuberculosis and sufficient to confer pathogenic properties to M. smegmatis.
研究领域:
微生物学
·
免疫学
·
致病机制
·
细胞生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
EUROPEAN JOURNAL OF PHARMACEUTICAL SCIENCES
·
2022
·
IF 5.112
Ping Zhu,Ling Cai,Qiao Liu,Shanwu Feng,Hongjie Ruan,Li Zhang,Liuzhu Zhou,Huijun Jiang,Hongsheng Wang,Jianming Wang,Jin Chen
Nowadays, pathogenic infection has posed a severe threat to the public health and environmental sanitation, urging a continuous search of efficacious and safe bactericidal agents of various formulated forms. Here, a facile one-pot hydrothermal preparation of mesoporous silica nanoparticles using ultrasonication-assisted nanoemulsion of α-Linolenic acid (α-LA) as template was developed. The formed silica mesocomposite at water/fatty-acid surface provides an easy yet green synthesis route, which can be generalized for the further encapsulation of hydrophobic drugs such as antimycobacterial Rifampicin (RIF). The obtained α-LA nanoemulsion-templated silica nanoparticles (LNS NPs), with a weight content of ∼17% α-LA in the composite, showed apparent antibacterial effect against Staphylococcus aureus ( S. aureus ). By comparison, the removal of α-LA from the silica nanoparticles (LNS-1 NPs) resulted in the composite of enlarged pore size with negligible bactericidal activities. Notably, the Isoniazide (INH) and Rifampicin (RIF)-encapsulated LNS NPs exhibited outstanding antimycobacterial activity against both drug-sensitive and drug-resistant Mycobacterium tuberculosis ( M. tuberculosis ). The obtained highly biocompatible, biosafe and low-energy consumptive α-LA-contained mesostructured silica-based bactericide holds promising therapeutic potentials to tackle the emerging drug-resistant infectious microbes.
研究领域:
材料科学
·
纳米技术
·
药物科学
·
细菌学
·
药物递送系统
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Current Drug Delivery
·
2023
·Vol. 20
·期 3
·306-316
·
IF 2.4
Jiang Huixian,Li Xiang,Xing Zhenjian,Niu Qun,Xu Jiangping
Background: Multidrug-resistant Mycobacterium tuberculosis (MDR-TB) is a major cause of death amongst tuberculosis patients. Nanomedicine avoids some limitations of conventional drug treatment and increases therapeutic efficacy against bacterial infections. However, the effect of anti-TB
drug nanoparticle (NP) compounds in anti-TB regimens against MDR-TB remains unclear. Objective: The objective of this article is to prepare levofloxacin, linezolid, ethambutol, prothionamide, and pyrazinamide encapsulated NPs and to evaluate their therapeutic efficacy against MDR-TB in
macrophages. Methods: Drug-loaded PLGA NPs were prepared by the multiple emulsion method. The colocalization, intracellular release, and anti-TB activity of these NPs were investigated on cultured macrophages. The immune phenotype of the macrophages, including their mitochondrial membrane
potential, reactive oxygen species (ROS), and nitric oxide (NO) production, was evaluated following treatment with NPs or free drug compounds. Results: All drug-loaded PLGA NPs were spherical in shape, 150 to 210 nm in size, and showed 14.22% to 43.51% encapsulation efficiencies and long-duration
release. Drug-loaded PLGA NPs were mainly distributed in the cytoplasm of macrophages, showed high cellular compatibility, and maintained their concentration for at least 13 days. Compared with the free drug compounds, the number of colonies after exposure to PLGA NP compounds was significantly
less. The enhanced antibacterial activity of the NP compounds may be due to the enhanced levels of ROS and NO and the increased early apoptosis stress within M. tuberculosis-infected macrophages additionally. Conclusion: The application of PLGA NP compounds not only enhances drug efficacy
but also induces innate bactericidal events in macrophages, confirming this as a promising approach for MDR-TB therapy.
研究领域:
药理学
·
微生物学
·
纳米医学
·
传染病学
✓ 已验证
DOI
PubMed
高影响因子
METABOLIC ENGINEERING
·
2019
·
IF 7.808
Wan-Ju Sun,Li Wang,Hao-Hao Liu,Yong-Jun Liu,Yu-Hong Ren,Feng-Qing Wang,Dong-Zhi Wei
The conversion of sterols to steroid synthons by engineered mycobacteria comprises one of the basic ways for the production of steroid medications in the pharmaceutical industry. Here, we revealed that high amounts of reactive oxygen species (ROS) generate during the conversion process of sterols, which impairs the cell viability of mycobacterial cells and thus hinders the conversion of sterols to steroid synthons. Accordingly, the endogenous antioxidants for detoxifying ROS in mycobacteria, ROS scavenging enzymes and low molecular weight thiols, were examined. The results revealed that three antioxidants, catalase (CAT), mycothiol (MSH), and ergothioneine (EGT), demonstrated efficacy toward neutralizing the excessive ROS produced during sterol metabolism . CAT overexpression or MSH or EGT augmentation enhanced the conversion of phytosterols to 22-hydroxy-23,24-bisnorchol-4-ene-3-one (4-HBC) by 18.9%, 23.8%, and 32.1%, respectively, and also enhanced the cell viability, indicating the benefits of these antioxidants in reducing ROS-induced stress. Further combinatorial augmentation of CAT, MSH, and EGT demonstrated enhanced effects toward intracellular ROS scavenging, resulting in 54.2% greater cell viability and 47.5% enhancement in 4-HBC production. These findings indicated that the excessive ROS induces cell stress, in turn limiting the conversion of sterols, whereas neutralization of the excessive ROS by combined control of CAT, MSH, and EGT serves as an effective strategy to boost the conversion productivity of sterols to steroid synthons.
研究领域:
药学
·
微生物学
·
生物技术
·
生物化学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
PLoS One
·
2014
·Vol. 9
·期 4
·e94418
·
IF 3.534
Wu Li,Quanju Zhao,Wanyan Deng,Tian Chen,Minqiang Liu,Jianping Xie
Intracellular survival plays a central role in the pathogenesis of Mycobacterium tuberculosis , a process which depends on an array of virulence factors to colonize and replicate within the host. The M. tuberculosis iron regulated open reading frame (ORF) rv3402c , encoding a conserved hypothetical protein, was shown to be up-regulated upon infection in both human and mice macrophages. To explore the function of this ORF, we heterologously expressed the rv3402c gene in the non-pathogenic fast-growing Mycobacterium smegmatis strain, and demonstrated that Rv3402c, a cell envelope-associated protein, was able to enhance the intracellular survival of recombinant M. smegmatis . Enhanced growth was not found to be the result of an increased resistance to intracellular stresses, as growth of the Rv3402c expressing strain was unaffected by iron depletion, H 2 O 2 exposure, or acidic conditions. Colonization of macrophages by M. smegmatis expressing Rv3402c was associated with substantial cell death and significantly greater amount of TNF-α and IL-1β compared with controls. Rv3402c-induced TNF-α and IL-1β production was found to be mediated by NF-κB, ERK and p38 pathway in macrophages. In summary, our study suggests that Rv3402c delivered in a live M. smegmatis vehicle can modify the cytokines profile of macrophage, promote host cell death and enhance the persistence of mycobacterium within host cells.
研究领域:
微生物学
·
免疫学
✓ 已验证
DOI
高影响因子
Frontiers of Environmental Science & Engineering
·
2019
·Vol. 14
·期 1
·1-11
·
IF 3.883
Wu Yiquan,Xu Ying,Zhou Ningyi
NidA3B3 is a terminal dioxygenase whose favorable substrates are high-molecular-weight polyaromatic hydrocarbons (PAHs) from Mycobacterium vanbaalenii PYR-1, a powerful PAHs degradation strain. NidA3B3 was reported to incorporate a dioxygen into the benzene ring of PAHs when equipped with an exogenous electron transport chain components PhdCD from Nocardioides sp. strain KP7 by biotransformation, but this enzyme system was not particularly efficient. In this study, strain PYR-1 was confirmed to utilize four different PAHs at different growth rates. When PhtAcAd, an endogenous electron transport chain ofa phthalate dioxygenase system, was substituted for PhdCD to couple with NidA3B3, the specific activity to convert phenanthrene by strain BL21(DE3) [pNidA3B3-PhAcAd] was 0.15±0.03 U/mg, but the specificactivity of strain BL21(DE3) [pNidA3B3-PhdCD] was only 0.025±0.006 U/mg. In addition, FNidA3, encoded by a newly defined ORF, has a prolonged 19-amino acid sequence at the N-terminus compared with NidA3. FNidA3B3 increased the activity by 50% approximately than NidA3B3 when using PhtAcAd. Components of the electron transport chain PhtAc and PhtAd were purified and characterized. The K m , k cat , k cat / K m values of the PhtAd were 123±26.9 µM, 503±49.9 min −1 , 4.1 µM −1 ×min −1 , respectively. And the; K m , k cat , k cat / K m values of the ferredoxin PhtAc were 52.5±9.7 µM, 3.8±0.19 min −1 and 0.07 µM −1 ×min −1 respectively. Basing on the phylogenetic analysis, NidA3/FNidA3 were far from its isoenzyme NidA from the same strain. Combining their primary differences of transcriptional pattern in vivo, it indicated that the functionally similar Rieske dioxygenases NidA3B3/FNidA3B3 and NidAB might originate from different ancestors.
研究领域:
微生物学
·
生物化学
·
环境科学
✓ 已验证
DOI
高影响因子
Frontiers in Cellular and Infection Microbiology
·
2020
·
IF 4.123
Ali Md Kaisar,Zhen Gong,Nzungize Lambert,Stojkoska Andrea,Duan Xiangke,Li Chunyan,Duan Wei,Xu Junqi,Xie Jianping
The Mycobacterium (M.) tuberculosis comprising proline–glutamic acid (PE) subfamily proteins associate with virulence, pathogenesis, and host-immune modulations. While the functions of most of this family members are not yet explored. Here, we explore the functions of “PE only” subfamily member PE31 (Rv3477) in virulence and host-pathogen interactions. We have expressed the M. tuberculosis PE31 in non-pathogenic Mycobacterium smegmatis strain (Ms_PE31) and demonstrated that PE31 significantly altered the cell facet features including colony morphology and biofilm formation. PE31 expressing M. smegmatis showed more resistant to the low pH, diamide, H2O2 and surface stress. Moreover, Ms_PE31 showed higher intracellular survival in macrophage THP-1 cells. Ms_PE31 significantly down-regulated the production of IL-12p40 and IL-6, while up-regulates the production of IL-10 in macrophages. Ms_PE31 also induced the expression of guanylate-binding protein-1 (GBP-1) in macrophages. Further analysis demonstrates that Ms_PE31 inhibits the caspase-3 activation and reduces the macrophages apoptosis. Besides, the NF-κB signaling pathway involves the interplay between Ms_PE31 and macrophages. Collectively, our finding identified that PE31 act as a functionally relevant virulence factor of M. tuberculosis.
研究领域:
微生物学
·
免疫学
·
病原学
·
宿主-病原体相互作用
·
细胞生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）