[go: up one dir, main page]

CN103160571A - 磷酸化肌球蛋白和驱动蛋白的激酶的发现 - Google Patents

磷酸化肌球蛋白和驱动蛋白的激酶的发现 Download PDF

Info

Publication number
CN103160571A
CN103160571A CN2011104077969A CN201110407796A CN103160571A CN 103160571 A CN103160571 A CN 103160571A CN 2011104077969 A CN2011104077969 A CN 2011104077969A CN 201110407796 A CN201110407796 A CN 201110407796A CN 103160571 A CN103160571 A CN 103160571A
Authority
CN
China
Prior art keywords
unc
myosins
kinesins
kinesin
myosin
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.)
Pending
Application number
CN2011104077969A
Other languages
English (en)
Inventor
田怀泽
李素云
田华希
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.)
RAINBOW HEALTH LIFE SCIENCES INSTITUTE (BEIJING) Co Ltd
Original Assignee
RAINBOW HEALTH LIFE SCIENCES INSTITUTE (BEIJING) Co Ltd
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 RAINBOW HEALTH LIFE SCIENCES INSTITUTE (BEIJING) Co Ltd filed Critical RAINBOW HEALTH LIFE SCIENCES INSTITUTE (BEIJING) Co Ltd
Priority to CN2011104077969A priority Critical patent/CN103160571A/zh
Publication of CN103160571A publication Critical patent/CN103160571A/zh
Pending legal-status Critical Current

Links

Images

Landscapes

  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)
  • Enzymes And Modification Thereof (AREA)

Abstract

生命在于运动。肌球蛋白(myosin)和驱动蛋白(kinesin)作为细胞运动的主要分子马达，在肌肉收缩，细胞内物质，小胞和膜细胞器的输送以及动态细胞骨架的构成和功能上起着重要的作用。揭示调节肌球蛋白和驱动蛋白功能的基本分子原理是生命科学家的最热衷研究领域之一。还没有关于既磷酸化肌球蛋白也磷酸化驱动蛋白的激酶的报道。在这里，首次揭示肌球蛋白和驱动蛋白都能够结合UNC-51激酶，首次揭示UNC-51蛋白激酶活性既能够磷酸化肌球蛋白也能够磷酸化驱动蛋白，并提出UNC-51激酶活性通过对肌球蛋白和驱动蛋白的磷酸化反应诱导神经轴突形成的模型。

Description

磷酸化肌球蛋白和驱动蛋白的激酶的发现
技术领域：
本发明属于基础生命科学，生物化学，细胞生物学，分子生物学，脑科学，神经细胞骨架学和神经细胞信号转导学等多个领域，创新性地揭示了同时调节细胞肌球蛋白和驱动蛋白的新机理和神经细胞形成及工作的基本原理。 
背景技术：
生命在于运动，探索生命运动的原理是生命科学的最重要领域之一。肌球蛋白(myosin)和驱动蛋白(kinesin)作为细胞运动的主要分子马达，在肌肉收缩，细胞内物质，小胞和膜细胞器的输送以及动态细胞骨架的构成和功能上起着重要的作用。 
细胞微管和微丝的动态和神经细胞轴突的形成，细胞分裂以及细胞移动等有密切的关系。肌球蛋白和驱动蛋白分别与细胞微丝和细胞微管紧密地协作，担负着各种复杂细胞运动的使命。揭示调节肌球蛋白和驱动蛋白功能的基本分子原理是生命科学家的最热衷研究领域之一。还没有关于既磷酸化肌球蛋白也磷酸化驱动蛋白的激酶的报道。模式生物秀丽线虫和哺乳动物的神经系统在形态学和生物化学上是相似的；一个保存性基因在秀丽线虫和哺乳动物的不同实验系里能表达相同的功效。秀丽线虫的unc-51基因编码一个从线虫到人类的进化上具有保存性的丝/苏氨酸蛋白激酶，是神经形成所必须的；这个基因的突变引起线虫的神经形成及运动的欠缺和短粗体形。揭示unc-51基因所编码的蛋白激酶的底物和其生物化学功能的意义是重要的。 
发明内容：
本发明的目的在于揭示unc-51基因所编码的蛋白激酶的重要底物和其生物化学功能，弄清楚神经细胞形成的分子机理，为神经疾病的治疗提供崭新的原理和方法。本发明的内容为： 
(1)首次揭示肌球蛋白和驱动蛋白都能够结合UNC-51激酶 
肌球蛋白及驱动蛋白和UNC-51激酶的体外结合如图1所示。图1显示体外酶联免疫吸附测定(ELISA)的结果，肌球蛋白重链(myosin heavy chain，MHC)，肌球蛋白轻链(myosin light chain，MLC)和驱动蛋白重链(myosin heavy chain，KHC)都能够在体外结合UNC-51激酶。 
(2)首次揭示UNC-51蛋白激酶活性既能够磷酸化肌球蛋白也能够磷酸化驱动蛋白 
图2显示UNC-51激酶在体外能够磷酸化肌球蛋白重链(MHC)并自我磷酸化。 
图3显示UNC-51激酶在体外能够磷酸化肌球蛋白轻链(MLC)并自我磷酸化。 
图4显示用凝胶移位分析法的结果，UNC-51激酶在体内(HeLa细胞)能够磷酸化驱动蛋白。 
图5表示UNC-51激酶活性通过对肌球蛋白和驱动蛋白的磷酸化反应诱导神经轴突形成的模型。 
附图说明：
图1肌球蛋白及驱动蛋白和UNC-51激酶的体外结合 
图2UNC-51激酶在体外磷酸化微管蛋白并自我磷酸化 
图3UNC-51激酶在体外磷酸化微管蛋白并自我磷酸化 
图4UNC-51激酶在体外磷酸化微管蛋白并自我磷酸化 
图5UNC-51激酶活性通过对肌球蛋白及驱动蛋白的磷酸化反应诱导神经轴突形成的模型 
具体实施方式：
【1】肌球蛋白及驱动蛋白和UNC-51蛋白激酶结合的测定 
用细胞溶解缓冲液溶解表达UNC-51蛋白激酶的HEK293细胞，得到溶胞产物的上清。利用亲和层析法从以上溶胞产物的上清纯化得到UNC-51蛋白激酶。运用固相酶联免疫测定法(ELISA)分析肌球蛋白及驱动蛋白和UNC-51蛋白激酶的体外结合。 
【2】激酶活性的测定 
在含有50μM末端放射性表记的腺苷三磷酸[γ-32P]-ATP的Hepes激酶缓冲反应液(pH7.4)中，进行了UNC-51蛋白激酶磷酸化肌球蛋白及驱动蛋白的反应。用6.5％的SDS-PAGE和X-线胶片放射自显影法分析测定了激酶活性。 
用凝胶移位分析法发现了UNC-51激酶在体内(HeLa细胞)能够磷酸化驱动蛋白。 

Claims (3)

  1. 本发明首次发现第一个既结合肌球蛋白及驱动蛋白也磷酸化肌球蛋白及驱动蛋白的激酶是UNC-51激酶。其关键发现为：
    1.在试管内的生化实验中，首次揭示肌球蛋白(MHC和MLC)和驱动蛋白(KHC)都能够结合UNC-51激酶。
  2. 2.首次揭示UNC-51蛋白激酶活性既能够磷酸化肌球蛋白(MHC和MLC)也能够磷酸化驱动蛋白(KHC)。
    肌球蛋白+UNC-51+驱动蛋白+ATP(GTP)→
    →移动性p-肌球蛋白-UNC-51-驱动蛋白-p复合体+ADP(GDP)
  3. 3.上记二项要求的技术原理在生命科学，脑科学，肿瘤科学，计算机科学，宇宙科学，交通科学，医学，药学和制药学，中药科学，各种疾病的治疗法，医疗技术以及所有其他方面的应用和推广。
CN2011104077969A 2011-12-09 2011-12-09 磷酸化肌球蛋白和驱动蛋白的激酶的发现 Pending CN103160571A (zh)

Priority Applications (1)

Application Number Priority Date Filing Date Title
CN2011104077969A CN103160571A (zh) 2011-12-09 2011-12-09 磷酸化肌球蛋白和驱动蛋白的激酶的发现

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN2011104077969A CN103160571A (zh) 2011-12-09 2011-12-09 磷酸化肌球蛋白和驱动蛋白的激酶的发现

Publications (1)

Publication Number Publication Date
CN103160571A true CN103160571A (zh) 2013-06-19

Family

ID=48584105

Family Applications (1)

Application Number Title Priority Date Filing Date
CN2011104077969A Pending CN103160571A (zh) 2011-12-09 2011-12-09 磷酸化肌球蛋白和驱动蛋白的激酶的发现

Country Status (1)

Country Link
CN (1) CN103160571A (zh)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1168674A (zh) * 1994-10-14 1997-12-24 昆士兰医学研究所 合成肽及包含其的疫苗
WO2004113561A2 (en) * 2003-06-25 2004-12-29 Queen's University At Kingston Methods for diagnosing, monitoring, staging and treating heart failure
CN101616909A (zh) * 2006-12-27 2009-12-30 塞诺菲-安万特股份有限公司 作为Rho-激酶抑制剂的被取代的异喹啉和异喹啉酮衍生物

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN1168674A (zh) * 1994-10-14 1997-12-24 昆士兰医学研究所 合成肽及包含其的疫苗
WO2004113561A2 (en) * 2003-06-25 2004-12-29 Queen's University At Kingston Methods for diagnosing, monitoring, staging and treating heart failure
CN101616909A (zh) * 2006-12-27 2009-12-30 塞诺菲-安万特股份有限公司 作为Rho-激酶抑制剂的被取代的异喹啉和异喹啉酮衍生物

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
TIAN HUAIZE,ET AL.: "UNC-51 phosphorylates tubulin/kinesin and actin/myosin and regualtes dynamics of microtubule/kinesin and actin/myosin systems by a competition-coordination mechanism", 《生化学》, 31 December 2006 (2006-12-31) *
杜军等: "肌动蛋白磷酸化在溶血磷脂酸致乳腺癌细胞迁移中的作用", 《南京医科大学学报(自然科学版)》, vol. 29, no. 5, 31 May 2009 (2009-05-31), pages 609 - 612 *

Similar Documents

Publication Publication Date Title
Canty et al. Structure and mechanics of dynein motors
Zhang et al. Identification of the functional binding pocket for compounds targeting small-conductance Ca2+-activated potassium channels
Rostas et al. Calcium/calmodulin-stimulated protein kinase II (CaMKII): different functional outcomes from activation, depending on the cellular microenvironment
Weirich et al. Structure, activity and function of the Suv39h1 and Suv39h2 protein lysine methyltransferases
Liou et al. In silico analysis reveals sequential interactions and protein conformational changes during the binding of chemokine CXCL-8 to its receptor CXCR1
McCarty et al. Escherichia coli QueD is a 6-carboxy-5, 6, 7, 8-tetrahydropterin synthase
Manschwetus et al. A stapled peptide mimic of the pseudosubstrate inhibitor PKI inhibits protein kinase A
Moussouras et al. Differences in sulfotyrosine binding amongst CXCR1 and CXCR2 chemokine ligands
Kraljić et al. Benzoxaboroles—novel autotaxin inhibitors
Zhu et al. Recombinant expression and characterization of α-Conotoxin LvIA in Escherichia coli
Fujino et al. Chimeric RNA oligonucleotides incorporating triazole-linked trinucleotides: synthesis and function as mRNA in cell-free translation reactions
Eletskaya et al. Enzymatic synthesis of 2-chloropurine arabinonucleosides with chiral amino acid amides at the C6 position and an evaluation of antiproliferative activity in vitro
CN103160571A (zh) 磷酸化肌球蛋白和驱动蛋白的激酶的发现
Miao et al. Targeted proteomic approaches for proteome-wide characterizations of the AMP-binding capacities of kinases
Liu et al. Optimization of phage heptapeptide library-screening process for developing inhibitors of the isocitrate lyase homologue from Mycobacterium tuberculosis
Cheng Cyclic nucleotide signaling
Varano et al. Design and synthesis of novel thiazolo [5, 4-d] pyrimidine derivatives with high affinity for both the adenosine A1 and A2A receptors, and efficacy in animal models of depression
CN103160569A (zh) 调节细胞微丝动态新机制的发现
JP2022065113A (ja) ホスホグリセリン酸ムターゼのペプチド阻害剤および使用方法
CN103160572A (zh) 调节细胞微管动态新机制的发现
Horrell et al. What a twist: structural biology of the SARS-CoV-2 helicase nsp13
Heissler et al. Tools to study nonmuscle myosin-2 motor function revisited
El-Mowafi et al. Structural Analysis and Activity Correlation of Amphiphilic Cyclic Antimicrobial Peptides Derived from the [W4R4] Scaffold
Vrancken et al. The symmetric designer protein pizza as a scaffold for metal coordination
Schmidt Nanoparticle-induced membrane pore formation studied with lipid bilayer arrays

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
C02 Deemed withdrawal of patent application after publication (patent law 2001)
WD01 Invention patent application deemed withdrawn after publication

Application publication date: 20130619