通过氘化辅助小角中子散射揭示同寡聚蛋白亚基交换的动力学不对称性。
Kinetic asymmetry of subunit exchange of homooligomeric protein as revealed by deuteration-assisted small-angle neutron scattering.
机构信息
Research Reactor Institute, Kyoto University, Kumatori, Osaka, Japan.
出版信息
Biophys J. 2011 Oct 19;101(8):2037-42. doi: 10.1016/j.bpj.2011.09.004.
Abstract
We developed a novel, to our knowledge, technique for real-time monitoring of subunit exchange in homooligomeric proteins, using deuteration-assisted small-angle neutron scattering (SANS), and applied it to the tetradecamer of the proteasome α7 subunit. Isotopically normal and deuterated tetradecamers exhibited identical SANS profiles in 81% D(2)O solution. After mixing these solutions, the isotope sensitive SANS intensity in the low-q region gradually decreased, indicating subunit exchange, whereas the small-angle x-ray scattering profile remained unchanged confirming the structural integrity of the tetradecamer particles during the exchange. Kinetic analysis of zero-angle scattering intensity indicated that 1), only two of the 14 subunits were exchanged in each tetradecamer and 2), the exchange process involves at least two steps. This study underscores the usefulness of deuteration-assisted SANS, which can provide quantitative information not only on the molecular sizes and shapes of homooligomeric proteins, but also on their kinetic properties.
摘要
我们开发了一种新颖的、据我们所知的技术，用于实时监测同寡聚体蛋白质中的亚基交换，使用氘辅助小角中子散射（SANS），并将其应用于蛋白酶体 α7 亚基的十四聚体。在 81% D2O 溶液中，同位素正常和氘代的十四聚体表现出相同的 SANS 图谱。混合这些溶液后，低 q 区域的同位素敏感 SANS 强度逐渐降低，表明亚基交换，而小角 X 射线散射谱保持不变，证实了交换过程中十四聚体颗粒的结构完整性。零角散射强度的动力学分析表明，1）每个十四聚体中只有两个亚基被交换，2）交换过程至少涉及两个步骤。这项研究强调了氘辅助 SANS 的有用性，它不仅可以提供同寡聚体蛋白质的分子大小和形状的定量信息，还可以提供其动力学性质的定量信息。
相似文献
1
Kinetic asymmetry of subunit exchange of homooligomeric protein as revealed by deuteration-assisted small-angle neutron scattering.通过氘化辅助小角中子散射揭示同寡聚蛋白亚基交换的动力学不对称性。
Biophys J. 2011 Oct 19;101(8):2037-42. doi: 10.1016/j.bpj.2011.09.004.
2
Small-Angle Neutron Scattering of RNA-Protein Complexes.RNA-蛋白质复合物的小角中子散射。
Methods Mol Biol. 2020;2113:165-188. doi: 10.1007/978-1-0716-0278-2_13.
3
Selective deuteration of tryptophan and methionine residues in maltose binding protein: a model system for neutron scattering.麦芽糖结合蛋白中色氨酸和甲硫氨酸残基的选择性氘代：用于中子散射的模型系统。
Eur Biophys J. 2008 Jul;37(6):815-22. doi: 10.1007/s00249-008-0280-5. Epub 2008 Feb 15.
4
Structural Information on Bacterial Amyloid and Amyloid-DNA Complex Obtained by Small-Angle Neutron or X-Ray Scattering.通过小角中子散射或小角X射线散射获得的细菌淀粉样蛋白及淀粉样蛋白-DNA复合物的结构信息
Methods Mol Biol. 2022;2538:75-93. doi: 10.1007/978-1-0716-2529-3_6.
5
Biomolecular Deuteration for Neutron Structural Biology and Dynamics.用于中子结构生物学与动力学的生物分子氘代
Methods Enzymol. 2016;566:113-57. doi: 10.1016/bs.mie.2015.11.001. Epub 2015 Dec 22.
6
Disassembly of the self-assembled, double-ring structure of proteasome α7 homo-tetradecamer by α6.蛋白酶体α7同型十四聚体的自组装双环结构被α6拆解。
Sci Rep. 2015 Dec 14;5:18167. doi: 10.1038/srep18167.
7
Hydrogen-Deuterium Exchange Dynamics of NISTmAb Measured by Small Angle Neutron Scattering.通过小角中子散射测量 NISTmAb 的氢氘交换动力学。
Mol Pharm. 2023 Dec 4;20(12):6358-6367. doi: 10.1021/acs.molpharmaceut.3c00751. Epub 2023 Nov 14.
8
Structural characterization of an intrinsically disordered protein complex using integrated small-angle neutron scattering and computing.利用集成小角中子散射和计算技术对无规卷曲蛋白复合物进行结构特征分析
Protein Sci. 2023 Oct;32(10):e4772. doi: 10.1002/pro.4772.
9
Matchout deuterium labelling of proteins for small-angle neutron scattering studies using prokaryotic and eukaryotic expression systems and high cell-density cultures.利用原核和真核表达系统以及高细胞密度培养进行用于小角中子散射研究的蛋白质氘代匹配。
Eur Biophys J. 2017 Jul;46(5):425-432. doi: 10.1007/s00249-016-1186-2. Epub 2016 Nov 14.
10
Biological small-angle neutron scattering: recent results and development.生物小角中子散射：最新结果与进展。
Acta Crystallogr D Struct Biol. 2018 Aug 1;74(Pt 8):715-726. doi: 10.1107/S2059798318005016. Epub 2018 Jul 17.
引用本文的文献
1
COG0523 proteins: a functionally diverse family of transition metal-regulated G3E P-loop GTP hydrolases from bacteria to man.COG0523 蛋白：从细菌到人，一类功能多样的过渡金属调控的 G3E P 环 GTP 水解酶家族。
Metallomics. 2021 Aug 13;13(8). doi: 10.1093/mtomcs/mfab046.
2
Structural Fluctuations of the Human Proteasome α7 Homo-Tetradecamer Double Ring Imply the Proteasomal α-Ring Assembly Mechanism.人蛋白酶体 α7 同源十四聚体双环结构的动态变化暗示了蛋白酶体 α 环组装机制。
Int J Mol Sci. 2021 Apr 26;22(9):4519. doi: 10.3390/ijms22094519.
3
Elucidation of the mechanism of subunit exchange in αB crystallin oligomers.阐明 αB 晶状体蛋白寡聚物中亚基交换的机制。
Sci Rep. 2021 Jan 28;11(1):2555. doi: 10.1038/s41598-021-82250-z.
4
Mutational and Combinatorial Control of Self-Assembling and Disassembling of Human Proteasome α Subunits.人类蛋白酶体α 亚基自组装和拆卸的突变和组合控制。
Int J Mol Sci. 2019 May 9;20(9):2308. doi: 10.3390/ijms20092308.
5
Molecular and Structural Basis of the Proteasome α Subunit Assembly Mechanism Mediated by the Proteasome-Assembling Chaperone PAC3-PAC4 Heterodimer.蛋白酶体α 亚基组装机制的分子和结构基础，由蛋白酶体组装伴侣 PAC3-PAC4 异二聚体介导。
Int J Mol Sci. 2019 May 7;20(9):2231. doi: 10.3390/ijms20092231.
6
Structural insights on the dynamics of proteasome formation.蛋白酶体形成动力学的结构见解
Biophys Rev. 2018 Apr;10(2):597-604. doi: 10.1007/s12551-017-0381-4. Epub 2017 Dec 14.
7
Two-step process for disassembly mechanism of proteasome α7 homo-tetradecamer by α6 revealed by high-speed atomic force microscopy.高速原子力显微镜揭示的蛋白酶体α7 同源十四聚体解组装机制的两步过程。
Sci Rep. 2017 Nov 13;7(1):15373. doi: 10.1038/s41598-017-15708-8.
8
Structural characterization of the circadian clock protein complex composed of KaiB and KaiC by inverse contrast-matching small-angle neutron scattering.通过逆对比匹配小角中子散射对由 KaiB 和 KaiC 组成的生物钟蛋白复合物的结构特征进行分析。
Sci Rep. 2016 Oct 18;6:35567. doi: 10.1038/srep35567.
9
New insight into the dynamical system of αB-crystallin oligomers.对 αB-晶体蛋白寡聚物动力学系统的新认识。
Sci Rep. 2016 Jul 6;6:29208. doi: 10.1038/srep29208.
10
Disassembly of the self-assembled, double-ring structure of proteasome α7 homo-tetradecamer by α6.蛋白酶体α7同型十四聚体的自组装双环结构被α6拆解。
Sci Rep. 2015 Dec 14;5:18167. doi: 10.1038/srep18167.
本文引用的文献
1
RDC-assisted modeling of symmetric protein homo-oligomers.RDC辅助的对称蛋白质同源寡聚体建模。
Protein Sci. 2008 May;17(5):899-907. doi: 10.1110/ps.073395108.
2
Small angle neutron and X-ray scattering in structural biology: recent examples from the literature.结构生物学中的小角中子散射和小角X射线散射：文献中的近期实例
Eur Biophys J. 2008 Jun;37(5):531-41. doi: 10.1007/s00249-008-0259-2. Epub 2008 Jan 23.
3
Cooperation of multiple chaperones required for the assembly of mammalian 20S proteasomes.哺乳动物20S蛋白酶体组装需要多种伴侣蛋白的协同作用。
Mol Cell. 2006 Dec 28;24(6):977-84. doi: 10.1016/j.molcel.2006.11.015.
4
Application of a Theory of Enzyme Specificity to Protein Synthesis.酶特异性理论在蛋白质合成中的应用。
Proc Natl Acad Sci U S A. 1958 Feb;44(2):98-104. doi: 10.1073/pnas.44.2.98.
5
A heterodimeric complex that promotes the assembly of mammalian 20S proteasomes.一种促进哺乳动物20S蛋白酶体组装的异二聚体复合物。
Nature. 2005 Oct 27;437(7063):1381-5. doi: 10.1038/nature04106.
6
Kinetic stabilization of an oligomeric protein under physiological conditions demonstrated by a lack of subunit exchange: implications for transthyretin amyloidosis.在生理条件下寡聚蛋白的动力学稳定性通过亚基交换的缺乏得以证明：对转甲状腺素蛋白淀粉样变性的影响。
Biochemistry. 2005 Jun 28;44(25):9265-74. doi: 10.1021/bi050352o.
7
Hydrogen exchange methods to study protein folding.用于研究蛋白质折叠的氢交换方法。
Methods. 2004 Sep;34(1):51-64. doi: 10.1016/j.ymeth.2004.03.005.
8
Hierarchical map of protein unfolding and refolding at thermal equilibrium revealed by wide-angle X-ray scattering.通过广角X射线散射揭示的热平衡下蛋白质解折叠和重折叠的层次图。
Biochemistry. 2004 Jul 20;43(28):9036-49. doi: 10.1021/bi0499664.
9
Mass spectrometric approaches using electrospray ionization charge states and hydrogen-deuterium exchange for determining protein structures and their conformational changes.利用电喷雾电离电荷状态和氢-氘交换的质谱方法来确定蛋白质结构及其构象变化。
Mol Cell Proteomics. 2004 Jan;3(1):10-23. doi: 10.1074/mcp.R300010-MCP200. Epub 2003 Nov 16.
10
Spontaneous subunit exchange and biochemical evidence for trans-autophosphorylation in a dimer of Escherichia coli histidine kinase (EnvZ).大肠杆菌组氨酸激酶（EnvZ）二聚体中自发的亚基交换及反式自磷酸化的生化证据。
J Mol Biol. 2003 Jun 6;329(3):495-503. doi: 10.1016/s0022-2836(03)00446-7.
Zotero 插件