CN106039286A - 一种抗结核分枝杆菌感染的蛋白trim27 - Google Patents

一种抗结核分枝杆菌感染的蛋白trim27 Download PDF

Info

Publication number
CN106039286A
CN106039286A CN201610381995.XA CN201610381995A CN106039286A CN 106039286 A CN106039286 A CN 106039286A CN 201610381995 A CN201610381995 A CN 201610381995A CN 106039286 A CN106039286 A CN 106039286A
Authority
CN
China
Prior art keywords
trim27
aminoacid
aminoacid sequence
resistant
analog
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
CN201610381995.XA
Other languages
English (en)
Other versions
CN106039286B (zh
Inventor
刘翠华
汪静
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Institute of Microbiology of CAS
Original Assignee
Institute of Microbiology of CAS
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Institute of Microbiology of CAS filed Critical Institute of Microbiology of CAS
Priority to CN201610381995.XA priority Critical patent/CN106039286B/zh
Publication of CN106039286A publication Critical patent/CN106039286A/zh
Application granted granted Critical
Publication of CN106039286B publication Critical patent/CN106039286B/zh
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K38/00—Medicinal preparations containing peptides
    • A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • A61K38/1703—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates
    • A61K38/1709—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans from vertebrates from mammals
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
    • G01N2333/195—Assays involving biological materials from specific organisms or of a specific nature from bacteria
    • G01N2333/35—Assays involving biological materials from specific organisms or of a specific nature from bacteria from Mycobacteriaceae (F)
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N2500/00—Screening for compounds of potential therapeutic value

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Chemical & Material Sciences (AREA)
  • Immunology (AREA)
  • General Health & Medical Sciences (AREA)
  • Urology & Nephrology (AREA)
  • Molecular Biology (AREA)
  • Hematology (AREA)
  • Biomedical Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Veterinary Medicine (AREA)
  • Marine Sciences & Fisheries (AREA)
  • Epidemiology (AREA)
  • Public Health (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biotechnology (AREA)
  • Cell Biology (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Microbiology (AREA)
  • Zoology (AREA)
  • Animal Behavior & Ethology (AREA)
  • Food Science & Technology (AREA)
  • Physics & Mathematics (AREA)
  • Analytical Chemistry (AREA)
  • Biochemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Pathology (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

本发明公开了一种抗结核分枝杆菌感染的蛋白TRIM27，属于细胞生物学领域。本发明提供的抗结核分枝杆菌感染的蛋白TRIM27，可以用于开发治疗肺结核的药物，或者JNK/p38信号通路激活剂，或者NF‑κB信号通路抑制剂，为新型抗结核药物的研究提供新的思路和分子靶标，尤其在抗结核等多种药物的开发方面将有着广阔的前景。

Description

一种抗结核分枝杆菌感染的蛋白TRIM27
技术领域
本发明涉及一种抗结核分枝杆菌感染的蛋白TRIM27，属于细胞生物学领域。
背景技术
20世纪90年代，由于耐多药结核病(mul-tidrug-resistant tuberculosis，MDR-TB)和广泛耐药结核病(Extensively Drug-Resistant Tuberculosis，XDR TB)的出现和全球性播散，使全球结核病疫情出现回升势头。目前MDR-TB的诊断和治愈率低，同时相当一部分MDR-TB患者具有很强的传染性，因此全球的MDR-TB防控工作正面临严峻挑战。结核病原菌结核分枝杆菌(M.tuberculosis,Mtb)是一种典型的胞内菌，它进化出了高效复杂的逃避或干扰宿主免疫信号通路的方法，只有深入探究结核分枝杆菌胞内存活的分子机制才能为抗结核药物的研发提供新的作用靶点。
TRIM27蛋白是TRIM家族中的一员，它拥有RING finger、B-box和coiled-coil三个保守结构域。该蛋白在细胞质和细胞核中都有定位，其中定位于细胞核的TRIM27蛋白常作为转录抑制因子抑制目的基因转录。TRIM27蛋白也可在细胞质中起作用，比如作为E3连接酶介导底物泛素化，进而抑制其下游信号通路等。TRIM27蛋白在分枝杆菌感染宿主过程中发挥怎样的作用，目前尚不清楚。
发明内容
本发明首先提供了一种抗结核分枝杆菌感染药物，其有效成分是以下(a)或(b)或(c)：
(a)SEQ ID NO：1所示的氨基酸序列；
(b)在(a)中的氨基酸序列经过取代、缺失或添加一个氨基酸或几个氨基酸且具有抗病原菌胞内存活活性的由(a)衍生的多肽或其类似物；
(c)与(a)、(b)中氨基酸序列整体相似度在85％以上的由(a)衍生的多肽或其类似物。
本发明通过实验证明：(1)TRIM27能够抑制病原菌结核分枝杆菌的胞内存活。CFU结果显示，当敲除巨噬细胞U937中TRIM27的编码基因后，结核分枝杆菌在巨噬细胞U937胞内的存活能力增强；而且，正常的巨噬细胞U937中TRIM27的蛋白水平变化趋势与胞内结核分枝杆菌菌数一致，即当病原菌感染宿主细胞后，胞内TRIM27蛋白水平显著增强，随着菌体被宿主清除，胞内的TRIM27的蛋白水平也逐渐下降。可见：当病原菌感染宿主细胞后，TRIM27蛋白作为宿主限制因子发挥抗感染作用。(2)通过双荧光素酶报告系统及WesternBlotting实验发现在分枝杆菌感染巨噬细胞过程中，TRIM27能够显著促进炎性细胞因子的分泌，同时促进细胞凋亡，进而达到抗病原菌感染的目的。(3)进一步，研究结核分枝杆菌能够潜伏存活的分子机制，发现结核分枝杆菌中的效应蛋白PtpA会以TRIM27为靶点，抑制其发挥抗感染作用，抑制TRIM27介导的天然免疫信号通路的活化及细胞凋亡，促进病原菌的胞内存活，从而实现免疫逃逸。
本发明成果可以用于开发治疗肺结核的药物，或者JNK/p38信号通路激活剂，或者NF-κB信号通路抑制剂，为新型抗结核药物的研究提供新的思路和分子靶标，尤其在抗结核等多种药物的开发方面将有着广阔的前景。此外，本发明成果还对寻找新的抗菌药物靶点和新药筛选具有重要的理论意义。
附图说明
图1：TRIM27抑制分枝杆菌胞内存活
图2：TRIM27促进宿主炎性细胞因子的分泌
图3：TRIM27促进细胞凋亡
图4：TRIM27与结核分枝杆菌分泌蛋白PtpA相互作用
图5：PtpA抑制TRIM27介导的JNK信号通路的活化及细胞凋亡。
具体实施方式
实施例1TRIM27蛋白抗分枝杆菌感染
(A)实验方法：用耻垢分枝杆菌(M.smegmatis)和卡介苗(BCG)分别感染野生型和敲除TRIM27的巨噬细胞U937，通过对比不同时间点两种巨噬细胞内分枝杆菌CFU变化来证明TRIM27在抗病原菌感染过程中发挥作用。
(B)结果：耻垢分枝杆菌M.smegmatis感染巨噬细胞U937细胞2小时后CFU数最高，随着感染时间延长，分枝杆菌被逐渐清除(图1B)。BCG感染U937后CFU数在8小时内逐渐下降，24小时后CFU数开始上升，原因是BCG中含有很多与结核分枝杆菌相同的效应蛋白开始发挥干扰宿主天然免疫反应的作用，促进BCG的胞内存活(图1A)。对比野生型和TRIM27敲除型U937细胞中的CFU差异，结果表明TRIM27正常表达的U937细胞中分枝杆菌清除速度更快，而TRIM27敲除的U937细胞中分枝杆菌存活能力增强，即不同感染时间点的CFU数均显著高于野生型细胞。
实施例2TRIM27促进炎症细胞因子的分泌
(A)实验方法：用M.smegmatis(图2C,D)和BCG(图2A,B)分别感染野生型和敲除TRIM27的巨噬细胞U937，通过荧光实时定量PCR检测感染不同时间点细胞因子il8和il1b的基因转录水平变化。
(B)结果：对比野生型和TRIM27敲除型U937细胞中il8和il1b的基因转录水平在不同感染时间点的变化，结果表明在M.smegmatis和BCG感染过程中，TRIM27干扰的U937细胞中il8和il1b的基因转录水平都明显降低。因此，在分枝杆菌感染巨噬细胞过程中，TRIM27在促进炎性细胞因子的分泌抵抗病原菌感染方面发挥重要作用。
实施例3TRIM27促进病原菌感染细胞的细胞凋亡过程
(A)实验方法：用M.smegmatis分别感染野生型和TRIM27敲除的巨噬细胞，通过Western blotting检测感染不同时间点凋亡标志蛋白Caspase 3剪切活化程度(Cleaved-caspase 3)变化。
(B)结果：对比不同感染时间点野生型U937和TRIM27敲除的U937两种细胞中Cleaved-caspase 3蛋白水平，结果表明TRIM27敲除的U937细胞中Cleaved-caspase 3水平更低，即TRIM27对细胞凋亡有促进作用(图3)。
实施例4Mtb PtpA抑制TRIM27介导的JNK信号通路活化及细胞凋亡
(A)实验方法：用野生型M.smegmatis和过表达PtpA的M.smegmatis分别感染野生型U937细胞和TRIM27敲除的U937细胞，分析不同感染时间点JNK、p38的磷酸化水平变化以及Cleaved-caspase 3水平变化。
(B)结果：首先通过酵母双杂交实验证实结核分枝杆菌分泌蛋白PtpA与宿主TRIM27蛋白相互作用(图4)。进一步对比不同感染时间点细胞中JNK的磷酸化水平以及Cleaved-caspase 3水平，结果表明PtpA对JNK的磷酸化激活和细胞凋亡都有抑制作用(图5A)，而当TRIM27敲除后，对JNK的磷酸化和细胞凋亡的抑制作用都变弱了，该结果说明PtpA通过与TRIM27相互作用抑制其介导的JNK信号通路活化及细胞凋亡过程(图5B)。
虽然本发明已以较佳实施例公开如上，但其并非用以限定本发明，任何熟悉此技术的人，在不脱离本发明的精神和范围内，都可做各种的改动与修饰，因此本发明的保护范围应该以权利要求书所界定的为准。

Claims (5)

1.一种抗结核分枝杆菌感染药物，其特征在于，其有效成分是以下(a)或(b)或(c)：
(a)SEQ ID NO：1所示的氨基酸序列；
(b)在(a)中的氨基酸序列经过取代、缺失或添加一个氨基酸或几个氨基酸且具有抗病原菌胞内存活活性的由(a)衍生的多肽或其类似物；
(c)与(a)、(b)中氨基酸序列整体相似度在85％以上的由(a)衍生的多肽或其类似物。
2.一种治疗肺结核的药物，其特征在于，其有效成分是以下(a)或(b)或(c)：
(a)SEQ ID NO：1所示的氨基酸序列；
(b)在(a)中的氨基酸序列经过取代、缺失或添加一个氨基酸或几个氨基酸且具有抗病原菌胞内存活活性的由(a)衍生的多肽或其类似物；
(c)与(a)、(b)中氨基酸序列整体相似度在85％以上的由(a)衍生的多肽或其类似物。
3.一种JNK/p38信号通路激活剂，其特征在于，其有效成分是以下(a)或(b)或(c)：
(a)SEQ ID NO：1所示的氨基酸序列；
(b)在(a)中的氨基酸序列经过取代、缺失或添加一个氨基酸或几个氨基酸且具有抗病原菌胞内存活活性的由(a)衍生的多肽或其类似物；
(c)与(a)、(b)中氨基酸序列整体相似度在85％以上的由(a)衍生的多肽或其类似物。
4.一种NF-κB信号通路抑制剂，其特征在于，其有效成分是以下(a)或(b)或(c)：
(a)SEQ ID NO：1所示的氨基酸序列；
(b)在(a)中的氨基酸序列经过取代、缺失或添加一个氨基酸或几个氨基酸且具有抗病原菌胞内存活活性的由(a)衍生的多肽或其类似物；
(c)与(a)、(b)中氨基酸序列整体相似度在85％以上的由(a)衍生的多肽或其类似物。
5.TRIM27蛋白在筛选和开发治疗结核分枝杆菌感染的药物中的应用。
CN201610381995.XA 2016-06-01 2016-06-01 一种抗结核分枝杆菌感染的蛋白trim27 Active CN106039286B (zh)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN201610381995.XA CN106039286B (zh) 2016-06-01 2016-06-01 一种抗结核分枝杆菌感染的蛋白trim27

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201610381995.XA CN106039286B (zh) 2016-06-01 2016-06-01 一种抗结核分枝杆菌感染的蛋白trim27

Publications (2)

Publication Number Publication Date
CN106039286A true CN106039286A (zh) 2016-10-26
CN106039286B CN106039286B (zh) 2019-09-17

Family

ID=57172684

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201610381995.XA Active CN106039286B (zh) 2016-06-01 2016-06-01 一种抗结核分枝杆菌感染的蛋白trim27

Country Status (1)

Country Link
CN (1) CN106039286B (zh)

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107648597A (zh) * 2017-09-28 2018-02-02 中国科学院微生物研究所 一种ERK和JNK信号通路抑制剂Mce2E
CN109536470A (zh) * 2018-11-28 2019-03-29 中国科学院微生物研究所 一种抗结核分枝杆菌的靶点及其应用
WO2022141941A1 (zh) * 2020-12-30 2022-07-07 广东省科学院微生物研究所(广东省微生物分析检测中心) 含有特异性分子靶标的克罗诺杆菌标准菌株及其检测和应用

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2005081911A2 (en) * 2004-02-25 2005-09-09 Dana-Farber Cancer Institute, Inc. METHODS AND COMPOSITIONS FOR THE TREATMENT AND PREVENTION OF HIV INFECTION USING TRIM5α
EP2322224A1 (en) * 2008-09-01 2011-05-18 National University Corporation Nagoya University Potentiator of activity of anti-cancer agent and use thereof, and biomarker for prediction of prognosis in cancer patient and use thereof

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2005081911A2 (en) * 2004-02-25 2005-09-09 Dana-Farber Cancer Institute, Inc. METHODS AND COMPOSITIONS FOR THE TREATMENT AND PREVENTION OF HIV INFECTION USING TRIM5α
EP2322224A1 (en) * 2008-09-01 2011-05-18 National University Corporation Nagoya University Potentiator of activity of anti-cancer agent and use thereof, and biomarker for prediction of prognosis in cancer patient and use thereof

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
NP_006501: "zinc finger protein RFP [Homo sapiens]", 《GENBANK数据库》 *
QINGLIANG ZHENG等: "Siglec1 suppresses antiviral innate immune response by inducing TBK1 degradation via the ubiquitin ligase TRIM27", 《CELL RESEARCH》 *

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN107648597A (zh) * 2017-09-28 2018-02-02 中国科学院微生物研究所 一种ERK和JNK信号通路抑制剂Mce2E
CN109536470A (zh) * 2018-11-28 2019-03-29 中国科学院微生物研究所 一种抗结核分枝杆菌的靶点及其应用
WO2022141941A1 (zh) * 2020-12-30 2022-07-07 广东省科学院微生物研究所(广东省微生物分析检测中心) 含有特异性分子靶标的克罗诺杆菌标准菌株及其检测和应用

Also Published As

Publication number Publication date
CN106039286B (zh) 2019-09-17

Similar Documents

Publication Publication Date Title
Rogerson et al. Expression levels of Mycobacterium tuberculosis antigen‐encoding genes versus production levels of antigen‐specific T cells during stationary level lung infection in mice
Kannika et al. Molecular serotyping, virulence gene profiling and pathogenicity of Streptococcus agalactiae isolated from tilapia farms in Thailand by multiplex PCR
Yeung et al. Comparative phenotypic, molecular, and virulence characterization of Vibrio parahaemolyticus O3: K6 isolates
Jurado et al. Lucio’s leprosy: a clinical and therapeutic challenge
CN106039286A (zh) 一种抗结核分枝杆菌感染的蛋白trim27
Stanford et al. Mycobacteria and their world
Tamassia et al. IFN-β expression is directly activated in human neutrophils transfected with plasmid DNA and is further increased via TLR-4–mediated signaling
Marino et al. Johne’s disease in cattle: an in vitro model to study early response to infection of Mycobacterium avium subsp. paratuberculosis using RNA-seq.
Kock et al. Orientia tsutsugamushi is highly susceptible to the RNA polymerase switch region inhibitor Corallopyronin A in vitro and in vivo
Abo-Kadoum et al. Mycobacterium tuberculosis PE17 (Rv1646) promotes host cell apoptosis via host chromatin remodeling mediated by reduced H3K9me3 occupancy
Phillips et al. Cytokine mRNA expression in Mycobacteriam ulcerans-infected human skin and correlation with local inflammatory response
Zhou et al. Listeria monocytogenes in the Chinese food system: strain characterization through partial actA sequencing and tissue-culture pathogenicity assays
Martins de Sousa et al. Acute immune response to Mycobacterium massiliense in C57BL/6 and BALB/c mice
Madden et al. Mycobacterium tuberculosis infection triggers epigenetic changes that are enriched in a type I IFN signature
Badewa et al. Chemokine receptor 5 and its ligands in the immune response to murine tuberculosis
Nayak et al. First report of powdery mildew on Lagenaria siceraria caused by Podosphaera xanthii in India.
Kalani et al. The gene expression level of IFN-γR1 and IFN-γR2 in a murine model treated with Toxoplasma gondii and its products
Ranjbar et al. Molecular characterization of epidemic isolates of Vibrio cholerae O1 by arbitrarily primed PCR (AP-PCR)
Asgharzadeh et al. Differentiation of modern and ancestral Mycobacterium tuberculosis in Northwest region of Iran by screening for the presence of TbD1
Yan et al. MicroRNA-20b carried by mesenchymal stem cell-derived extracellular vesicles protects alveolar epithelial type II cells from Mycobacterium tuberculosis infection in vitro
Huang et al. First report of Citrus depressa as a new natural host of Xanthomonas citri subsp. citri pathotype A in Taiwan.
Peng et al. Effects of cefodizime on chemokines of liver tissues in mice with immunological hepatic injury
Kurniawati et al. Single nucleotide polymorphism of eccb5 gene of mycobacterium tuberculosis complex isolates from suspected pulmonary Tb patients in Surabaya Indonesia
Fallah et al. The history of tuberculosis and bacillus Calmette–Guérin vaccine in Iran
Saeed et al. ‘Ubiquitin: There’s no quitting

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
GR01 Patent grant
GR01 Patent grant