On September 6, 2018, local time, Yan Ning's research team published an article entitled "Structure of human-sourced gated sodium ion channels Nav1.4 and beta1 complex" (Structure of the human-gated sodium channel Voltage Nav1.4 in vs. vs. beta1) published online in Science. It reveals the frozen electron mirror structure of the first personal source voltage gated sodium ion channel (sodium channel) Nav1.4 and its specific regulation of the sub-beta 1 complex, with a resolution of up to 3.2 e (0.32 nanometers), providing a reliable template for a deep understanding of its mechanism of action and the mechanism of disease-related mutations.
1945, British scientists Hodgkin and Huxley first detected current on the giant neurons of gun squid and recorded the resting potential and action potential for the first time;
sodium channel is responsible for the occurrence and propagation of action potentials and is the basis for electrical signals on all neurons and muscle cells.
Hodgkin and Huxley won the 1963 Nobel Prize in Physiology and Medicine.
1970s, Neher and Sakmann began recording the current of a single ion channel using diaphragm pliers, and Nell first recorded currents in a single sodium channel in 1980.
they won the Nobel Prize in Physiology and Medicine in 1991.
1984, Japanese scientist Shosaku Numa and others first cloned the gene for sodium passages in electric eels.
decades, renowned scientists, including Hille, Armstrong, Bezanilla, catero, and other famous scientists, have contributed greatly to the biophysical and electrophysiological studies of sodium channels.
sodium channel is made up of alpha sub-bases responsible for the sensory voltage and ion selection, and 1-2 beta subakis that regulate it.
in the human body, alpha subtype has 9 different subtypes, named Nav1.1-1.9, beta subtype has 4 subtypes, respectively named beta 1-beta 4.
alpha subkeytypically contains about 2000 amino acids, containing four cross-membrane domain I-IV with similar sequences, each containing six transmembrane helixes.
the first four trans-diaphragm spiralS1-S4 in each domain constitute a voltage receptor (VSD), while four sets of S5-S6 trans-membrane spirals together form the responsible ion permeable hole domain domain located in the center of the entire structure, connecting the two semi-membrane short spirals between the trans-membrane helix S5-S6 to support a small segment of the long-range sequence, forming a selection sieve responsible for achieving ion permeability specificity.
the short sequence between the third and fourth domains connected has been shown to be critical for the rapid inactivation of sodium channels, but the exact mechanism is not clear.
rapid inactivation mechanism allows the sodium channel to close quickly after the sensory stimulation, causing the action potential, and preventthe persistent discharge of neuromuscular cells.
at the same time sodium potassium pump reconstruction membrane potential, ready for the next action potential.
the rapid inactivation of the sodium channel is therefore essential for it to perform normal physiological function. The nine subtypes of the
sodium channel have tissue distribution specificity, and their abnormal inactivation or activation is associated with a variety of serious neurological, cardiovascular, and muscle system diseases, such as Nav1.1 and Nav1.5 each have more than 400 point mutations associated with epilepsy and arrhythmia, and Abnormal itys in Nav1.4 can cause periodic paralysis of muscle stiffness or hyperkalemia, and abnormal pain in Nav1.7 or Nav1.8.
therefore, sodium channel is an important pharmaceutical target, is the focus of many international famous pharmaceutical companies research object.
, sodium channels are the direct targets of a variety of animal toxins, including snake poison, scorpion poison, puffer toxin, spider toxin, etc.
after more than 70 years of research, electrophysiological research on the working mechanism of human-derived sodium channels has been extensive, but because obtaining even trace amounts of high-quality samples is extremely difficult, so human-derived sodium channel is one of the most challenging proteins in structural biology.
Yanning has been working on structural biology of sodium pathways since he led the lab independently in 2007, and in 2012 reported on the crystal structure of NavRh,1, a sodium channel in a class of marine bacteria.
but the sodium channel of bacteria and eukaryotic organisms has some of the most basic differences, such as different ion selectivity, lack of rapid inactivation mechanism, can not be identified by toxins and so on, so can not become an alternative protein for the study of eyreum channels.
, Yan Ning has been focusing on the structure and mechanism of the eukaryokarita channel for the past five years, finally reporting the first eukaryokarita channel atomic model in 2017, the 3.8 ee resolution electron mirror structure (2) named NavPaS from the U.S. cockroach, and the electroscopic structure (3) of the electron eel channel, which plays an important role in the history of sodium channel research, and the first time in the structure of the sodium interaction.
Although these two structures represent a huge leap forward in the study of the structure and mechanism of sodium channels, neither channel can perform functional research in the external expression system, thus limiting them to become model research proteins. After several years of
, Professor Yan Ning's team finally overcome sedative purification of protein expression and frozen sample preparation and other technical bottlenecks step by step, analyzed the electromirror structure of the human-derived skeletal sodium channel subtype Nav1.4 up to 3.2 E, and revealed for the first time the structure of the complete voltage perception domain, ion selection sieve, fast inactivation originals and other key functional units.
on this basis, they will be found in muscle stiffness and hyper-potassium blood type periodic paralysis patients found more than 50 single-point mutations one-to-one correspondence to the structure, and focus on the voltage perception and rapid inactivation of the mutations, thus providing an important molecular basis for understanding the mechanism of disease. the trans-membrane and extracellular region structures of nav1.4-beta 1 in the
(A) human source Nav channel nav 1-beta 1 are well analyzed; (B) Nav1.4-beta 1 has a large negative area from the outside of the cell; (C) Nav1.4-beta 1's central hole is open from the inside of the cell; and (D) calculates the radius of Nav1.4-beta-1's central hole.
looking back over the past decade, Yan Ning's research team led by Tsinghua University has analyzed the structure of the original nuclear voltage gated sodium ion channel RhNav (1), the structure of the first high-resolution voltage gated calcium ion channel complex Cav1.1 (4,5), the structure of the first eukaryokaritchannel NavPaS (2), the first sodium channel and the structure of the regulatory sub-base (3), the first animal toxin and sodium channel structure (6), and finally obtained the structure of the first individual source of sodium.
because Nav1.4 has a definite physiological function, has been through biophysics, electrophysiology and other means in a variety of expression systems extensive research, and now its precise structure analysis opened a new page in the study of sodium channel structure and mechanism, to the full interpretation of the sodium channel process of the most important step.
Professor Yan Ning, former lying at Tsinghua University's School of Life and a center for high-precision innovation in structural biology, is the author of the research paper.
Pan Xiaojing, a postdoctoral scholar at Tsinghua University School of Medicine, a distinguished scholar at the Center for High-Excellence in Structural Biology, Li Zhangqiang, a third-year doctoral student at CLS, Zhou Qiang, an associate researcher at the Medical College, Adnessa, Shen Waizong, a distinguished scholar at the High-Level Innovation Center of Structural Biology, and Wu Kun, a doctoral student at Xiao Bailong Laboratory of the School of Pharmacy, are the co-first authors.
The National Protein Science Center (Beijing) Tsinghua University Cryptoscope Platform and Tsinghua University's high-performance computing platform supported the data collection and data processing of the research, respectively, And Dr. Lei Jianlin of Tsinghua University's Cryoscopic Mirror Platform guided the data collection and received technical assistance from staff member Li Xiaomin.
The Xiongyu Laboratory of Tsinghua University and The Bo Haipeng Laboratory provide guidance and assistance for electrophysiological and molecular dynamic simulation.
Beijing High-Tech Innovation Center for Structural Biology (Tsinghua), Joint Center for Life Sciences (Tsinghua University), National Key Laboratory for Biofilm and Membrane Bioengineering, Ministry of Science and Technology and Fund Committee provided financial support for the research. Since joining the Department of Molecular Biology at Princeton University in the fall of 2017, Professor
Yanning has been supported by Professor Shirley M. Tilghman's special start-up funding.
Source: Tsinghua University.