研究成果:Using 1-D Spatially Resolved Silicon-Nanowire-Field-Effect-Transistor (SiNW-FET) Platform to Characterize Cell Membrane Proteins in Their Native Bilayer Environment.

 

建立分析膜蛋白的仿生膜檢測平台
Using 1-D Spatially Resolved Silicon-Nanowire-Field-Effect-Transistor (SiNW-FET) Platform to Characterize Cell Membrane Proteins in Their Native Bilayer Environment.

膜蛋白為目前主要藥物標靶和發展新穎治療方式的研究對象,能夠快速檢測出能與目標膜蛋白結合之配體可有助於新藥發展。現今用於檢測配體結合的主流方式為螢光免疫染色法,然而,螢光標記方法很有可能會改變生物分子交互作用,而造成不正確的檢測結果。本計劃利用場效電晶體技術,來發展可偵測膜蛋白和配體結合反應之無標記檢測法。我們選擇具有二維結構之石墨烯作為場效電晶體半導體材料,以方便和二維支撐式脂質雙層膜做結合。我們發現石墨烯表面高分子殘留物會干擾脂質膜形成,需用高溫淬火將之移除來使得脂質膜能容易形成。此外,我們也發現石墨烯上方脂質膜螢光幾乎會完全被石墨烯猝滅,無法以傳統螢光恢復法來驗證脂質膜的形成,因此本計畫開發一種新方法來測試脂質膜形成狀況,以找尋適合的成膜條件。我們更進一步發現緩衝液中的離子會逐漸吸附到石墨烯表面,而導致石墨烯的狄拉克點會在配體結合檢測期間隨時間顯著偏移。透過仔細控制微流體裝置中緩衝液添加量和流動持續時間,我們得以去除由離子吸附引起的效應。我們使用鏈黴親和素與具有生物素之脂質作為模型,來測試配體結合在本平台上產生的效應,結果顯示本平台的確能定量地檢測到配體與脂質膜上物質結合。將此計畫發展出的平台進一步與我們由大腸桿菌所得到之細胞膜囊泡做結合,將可使感興趣之膜蛋白在其天然脂質雙層環境中來進行無標記檢測。

Cell membranes are the gateway for the cells to send and receive signals. More than 60% of commercial drugs interact with membrane proteins to alter cellular biochemical pathways. Since membrane proteins are primary pharmaceutical targets, information about what ligands can bind to them is useful to facilitate drug discovery and development. Most of the current methods to identify the ligand binding to membrane proteins rely on immunodetection. However, immunodetection methods may alter biomolecular interactions. Here, we develop a label-free detection method based on field effect transistor (FET) technique, which can provide rapid and sensitive responses to the ligand binding. Supported lipid bilayers (SLBs) have been widely used as platforms for studying interactions between ligands and membrane proteins in vitro. To build a FET detection platform combined with SLBs, we chose graphene as the FET semiconductive material because the two-dimensionality is compatible with the planar bilayer platform and it has high sensitivity. We fabricated graphene field effect transistor (GFET) on silica substrates and investigated the formation of SLBs on the graphene/silica substrate. We discovered that the poly(methyl methacrylate) (PMMA) residues left on graphene surface can significantly influence the formation of SLBs on graphene and applying an extra annealing step to further clean the graphene surface is required for the robust SLB formation. In addition, because the fluorescent dyes in the SLB above graphene were almost completely quenched by graphene, which hinders us from using the conventional fluorescence recovery after photobleaching (FRAP) method to verify the formation of SLBs, we developed a modified FRAP method to examine whether SLBs successfully formed on the graphene/silica substrate. Furthermore, we discovered that ions in a regular buffer can gradually adsorb to the graphene surface and cause the Dirac point to shift significantly with time during our ligand binding detection. With careful control of the history of buffer addition and flow duration in a microfluidic device, we were able to eliminate the response caused by the ion adsorption and obtained the response only from the ligand binding. We used streptavidin-biotin-lipid as a model ligand-receptor to examine the GFET response to ligand binding on the SLB. Our results show that the SLB-GFET is capable to detect the streptavidin molecules binding onto the biotin-lipids embedded in the SLB, and the consistency between the electrical response and the fluorescent quantification result shows the quantitative detection capability of our GFET device. Incorporating this platform with the technique to obtain membrane vesicles from E. coli could allow us to study the interactions between potential drug candidates and interested membrane proteins in their native-like environment.

 

 

 

研究成果:泛素化於溶小體功能與品管的影響

 

泛素化於溶小體功能與品管的影響
Selective ubiquitination in lysophagy, lysosome maintenance, and lysosomal disorders

計畫成果摘要
泛素化被認為是控制受質被細胞自噬分解的關鍵。我們在先前的實驗已經發現,受損的溶小體會被泛素化,接著被細胞自噬蛋白辨識,最後進而被分解。但是負責將受損的溶小體泛素化的泛素蛋白酶目前尚未被發現。利用這個計畫的資助我們試圖利用APEX的方式來找出可能幫助受損的溶小體泛素化的泛素蛋白酶。目前我們已經找出幾個可能的泛素蛋白酶,所以將會在未來利用各種實驗方法驗證他們在受損的溶小體被泛素化中扮演的確切角色。

Project abstract
Ubiquitination has been implicated as a key signal for the selective degradation of substrates through autophagy. We have previously discovered that damaged lysosomes undergo selective ubiquitination, recruit autophagic adaptors, and are eventually turned over through that autophagic pathway (termed lysophagy). It however remains unknown what ubiquitin ligases mediate this degradation pathway. With the help of this grant and others we attempted to develop an APEX-based strategy to identify the ubiquitin ligases involved. Using this strategy we identified a panel of candidates that may be responsible for lysophagy. These candidates will be subjected to future testing for complete understanding of the lysophagy pathway.

 

 

研究成果:發展催化劑能在常溫常壓下有效能地轉換甲烷至甲醇

 

發展催化劑能在常溫常壓下有效能地轉換甲烷至甲醇
Develop a catalyst for catalytic conversion of methane into methanol efficiently under ambient conditions

The conversion of methane into methanol is very difficult chemistry requiring catalysts operating at elevated temperatures and pressures. The prospects of a future methanol economy depend on the successful development of robust and efficient catalysts that can accomplish this chemistry under mild conditions. In nature, the particulate methane monooxygenase (pMMO), a membrane-bound metalloenzyme, catalyzes methane oxidation with total selectivity and atom efficiency. Inspired by what we have learned about the pMMO over the past two decades, a tricopper complex [CuICuICuI(7-N-Etppz)]1+, where 7-N-Etppz stands for the organic ligand 3,30-(1,4-diazepane-1,4-diyl)bis[1-(4-ethylpiperazine-1-yl)propan-2-ol] based on the catalytic site of this enzyme has been developed and shown to mediate methane oxidation to methanol selectively. However, as a homogeneous catalyst, the catalytic efficiency and product yield are low because of the low solubility of methane in the solvent supporting the catalyst.

To overcome the problem of methane solubility in the catalytic system and to further improves the product yield. We re-formulated the tricopper cluster complex as a heterogeneous catalytic system assembled by immobilizing the tricopper cluster complex into mesoporous silica nanoparticles (MSNs) to take advantage of the “over solubility” of methane in liquids confined in nanoporous materials relative to the bulk solubility. For these gases, the interactions of the gas molecules with the nanoporous solid or with the solvent are significantly weaker than the interactions of the solvent molecules with the walls of the confining host framework. The stronger solvent–solid interactions create regions of low solvent density in the confined solvent, enhancing gas uptakes like CH4 and O2. The much higher solubility of methane within the pores of the mesoporous silica nanoparticles, as compared to the bulk solubility, led to very efficient turnover of the concentrated confined methane. Thus, the heterogeneous formulation exhibits dramatically higher catalytic efficiencies and turnover numbers, with commensurate improvements in chemical yields, offering the most proficient catalyst for the selective conversion of methane into methanol at room temperature developed to date. In this manner, we can drive the conversion of methane into methanol with hydrogen peroxide with a catalytic efficiency approaching 100%, as well as with high energy and atom economy under ambient methane pressures. The heterogeneous catalyst is robust and reuseable at least three reaction cycles. The samples are easily separated from the reaction mixture and dried under vacuum after each reaction cycle before performing the next run. This technology will lead to an economic process for methanol production that could have potential for further commercial applications.

 


 

直接轉化甲烷至甲醇是十分困難的化學反應,通常需要操作於高溫高壓的條件下而在未來甲醇經濟(是一種提議中未來的經濟形式,使用甲醇來代替現在廣泛使用的化石燃料來用作能量存儲,地面交通燃料,以及合成碳氫化合物的原料及其產品)的前景取決於是否能研發出強壯以及高效率的催化劑,能於常溫常壓下轉化甲烷為甲醇。然而在大自然中,普遍存在於嗜甲烷菌中的細胞質內膜上的微粒體型甲烷單氧化酵素(pMMO)卻可在常溫下高選擇性轉換甲烷為甲醇從過去二十年研究微粒體型甲烷單氧化酵素所學的知識並模擬其酵素活性中心的結構所合成出的三銅金屬錯合物[CuICuICuI(7-N-Etppz)]1+,可以於常溫常壓下高選擇性地執行甲烷至甲醇的反應然而這種昀相催化劑的催化效率以及甲醇的產率仍然十分低,最主要的原因是甲烷在有機溶劑中的溶解度十分低所致

為了改善甲烷氣體在催化系統中的溶解度,以近一步提高甲醇的產率,我們將三銅金屬錯合物負載於官能基化的中孔洞二氧化矽奈米球材料的垂直孔道中,利用”過度溶解度”的特性來大幅度提升被限制於奈米孔洞中的甲烷的溶解度在限制空間中,由於非極性的甲烷氣體分子與奈米孔洞內壁以及溶劑之間的作用力顯著的弱於溶劑分子與奈米孔洞內壁骨架間的作用力,而這溶劑與骨架間的強作用力造成許多區域的溶劑分子濃度較低但非極性氣體分子的濃度大幅提高(例如氧氣及甲烷)高濃度的甲烷分子貯存於二氧化矽奈米球的孔洞中,可以顯著的提升甲烷的催化效率以及轉換數,有效率的提升產物甲醇的產率,並有可能發展成為至今最有效率的高選擇性常溫常壓下轉換甲烷為甲醇的催化劑此異相催化劑甚至可以達到百分之百轉換甲烷為甲醇,有很高的能量與原子的經濟性此外此催化劑也可以重複使用至少三次催化反應此項技術, 將邁向更經濟的直接甲烷氧化為甲醇的工業製程並十分有潛力於商業應用

研究成果:TDP-43蛋白片段寡聚物之生化及生醫研究

 

室溫量子自旋電子與聲子的媒合
Marriage of quantum spintronics and phononics at room temperature

In this project, we have ultilized the various techniques and experties of all PIs and co-PIs to develope the new research area of thermal transport on non-magnetic and magnetic nanowires. For the growth of magnetic nanowire of Mn-SiGe, we established a new method to introduce Mn gas precursors MnCl2 during the VLS nanowire growth in CVD, such that Mn and Si atoms could be absorbed from the vapor phase by the Au-Si liquid catalysts and co-precipitated to form Si-Mn alloy nanowires. For the thermal transport probe, we have developed a new and sensitive probe based on the concept of thermal rectification. The thermal rectification in a pristine SiGe nanowire is shown in the inset of Fig. 1(a). The 4ω signals display a non-zero background which decreases with increasing Is,DC (or Ts,DC) as shown in Fig. 1(a). Regardless of the presence of the background, the absence of a peak at Is,DC=16μA indicates that there is no thermal rectification (R-1<0.2%). The result can be further verified by measuring 1ω and 2ω signals described above. As shown in Fig. 1(b), the 1ω/2ω signals vs. Ih,DC display a linear relation without a bump, which consistently indicate R-1<0.2% in the SiGe nanowire.

For the non-magnetic crystalline nanowire, Bi2-xSbxTe3 (BST) nanowires were grown directly from Bi0.5Sb1.5Te3 thin film by thermal annealing and the diameter dependence of thermoelectric properties of single BST NW has been in-situ studied in FIB trimming. The consequence implies the size and defect effects have more influence on phonon scattering than that of electron carriers in this diameter region. When the NW was further trimmed to 285 nm, both the electrical and thermal conductivities started a dramatic drop due to the formation of a large number of defects and the amorphous-like structure. The study not only provides the thoroughly understanding of the size and ion irradiation effects on the electrical and thermal transport properties but also provides a possible method to manipulate the physical properties in nanostructures for the extensive applications in engineering high ZT thermoelectric materials.

FIG. 1. (a) The signals at 4ω of a SiGe nanowire when gradually raising the Ts,DC of the sensor. The inset shows the SEM image of the SiGe nanowire anchored between a heater and a sensor. (b) The 1ω/2ω signals of the same sample when gradually raising the Th,DC of the heater. From the data and our analyses, we determine R-1<0.2%. (c), (d) The diameter dependent electrical, thermal conductivity, seebeck coefficient and figure of merit and of a FIB trimmed BST nanowire.

In the near future, we will further use this technique to explore the dynamic magnetism and the thermal properties of for the quasi-one dimensional magnetic nanowires.

 


 

本計劃中我們研究了摻雜錳元素於矽奈米線以將其變成磁性半導體的可能性,並探討了各種奈米線的熱傳導與熱整流性質。在生長磁性奈米線方面,我們提出了使用氣態MnCl2作為參與Au-Si液態催化反應以析出Si-Mn合金的方法。在熱傳導與熱整流性質的研究上,我們發明了利用4ω方法研究了諸多奈米線的熱整流性質。由於此新方法排除了以往量測熱功率的不確定性,因此比以前的方法更靈敏約十倍(如圖一a所示)。另外我們也開發了1ω/2ω的量測方法,並實驗證實這兩種方法得到一致的結果(如圖一b所示)。以圖一的矽鍺奈米線為例,我們的實驗結果確認其熱整流效應小於0.2%。這個實驗方法將對於介面與非對稱結構上的聲子與熱傳導研究有基礎的重要性。未來我們將利用本計劃所開發的技術與方法研究準一維磁性奈米線並探索基礎的動態磁學與熱傳導問題。

對於非磁性結晶納米線方面,Bi2-xSbxTe3(BST)奈米的成長以通過熱處理技術,直接從Bi0.5Sb1.5Te3薄膜生長,並且整合FIB修剪技術對單根BST NW的熱電性能的原位研究技術探究其物理特性與奈米線直徑之相依性。 結果顯示:尺寸和缺陷效應對聲子散射的影響比在該直徑區域中的電子載流子的影響更大,當NW修剪至285nm時,由於形成大量缺陷和非晶狀結構,電導率和熱導率開始顯著下降。 該研究不僅提供了對尺寸和離子輻射效應對電和熱傳輸性能的徹底理解,而且提供了操縱納米結構中的物理性質的可能方法,可廣泛應用於高ZT熱電材料。

圖一(a) 以4ω方法量測奈米線的熱整流性質。嵌入圖為懸空的矽鍺奈米線,兩端固定於微米熱傳導量測裝置的電子顯微鏡影像。(b)以1ω/2ω方法量測奈米線的熱整流性質。綜合圖一a與b的結果我們確認其熱整。(c), (d)電,熱導率,塞貝克係數和品質因數(ZT)於FIB修剪後BST納米線及其對直徑的相依性。

104年度「國立臺灣大學與中央研究院創新性合作計畫」核定計畫

國立臺灣大學與中央研究院創新性合作計畫

 104年度核定通過計畫

項次 計畫名稱 計畫主持人 計畫期間

1

室溫量子自旋電子與聲子的媒合
Marriage of quantum spintronics and phononics at room temperature
台大凝態中心-林昭吟
中研院物理所-陳洋元
104.7.1 ~
105.12.31

2

發展催化劑能在常溫常壓下有效能地轉換甲烷至甲醇 台大化學系-牟中原
中研院化學所-俞聖法
104.7.1 ~
105.12.31

3

Selective ubiquitination in lysophagy, lysosome maintenance, and lysosomal disorders 台大生科系-黃偉邦
中研院生化所-楊維元
104.7.1 ~
105.12.31

4

Using 1-D Spatially Resolved Silicon-Nanowire-Field-Effect-Transistor (SiNW-FET) Platform to Characterize Cell Membrane Proteins in Their Native Bilayer Environment. 台大化工系-趙玲
中研院生化所-史有伶
104.7.1 ~
105.12.31

5

以化學光遺傳小鼠模式探討止癢產生愉悅感的大腦神經基礎與酬賞機制神經迴路
Neurobiological basis of rewarding circuitry involved in itch relief-induced pleasure: a chemo-optogenetic approach
台大心理系-賴文崧
中研院生醫所-陳志成
104.7.1 ~
105.12.31

6

發展針對阿茲海默症中乙形類澱粉蛋白寡聚體之新穎小分子抑制物及核磁影像探針
Development of Novel Small Molecule Inhibitors and MRI Probes Targeting Alzheimer’s Amyloid-β Oligomers
台大化學系-詹益慈
中研院基因體中心-陳韻如
104.7.1 ~
105.12.31

7

鑑定克雷伯氏菌引發的‘嗜中性球胞外捕捉’及肝膿瘍的分子機制
Identification of the molecular mechanism of Klebsiella pneumoniae NTUH K2044-induced NETs formation and liver abscess
台大微生物學科-王錦堂
中研院基因體中心-謝世良
104.7.1 ~
105.12.31

8

肝病奈米組合藥物研發
Combination Nanotherapy of Liver Diseases
台大微生物學科-楊宏志
中研院生醫所-施嘉和
104.7.1 ~
105.12.31

9

C4與C3植物維管束鞘細胞及葉肉細胞發育之不同
Differences in bundle sheath and mesophyll cell development between C4 and C3 plants
台大植物所-鄭石通
中研院多樣中心-李文雄
104.7.1 ~
105.12.31

10

以hTERT基因探討DNA甲基化與G-quadruplex之交互影響基因表現機制 台大生化科技系-陳進庭
中研院原分所-張大釗
104.7.1 ~
105.12.31

11

Cell-type specific structural and functional studies of MD-InC thalamocortical neurons in neuropathic pain 台大生科系-陳示國
中研院生醫所-徐百川
104.7.1 ~
105.12.31

12

An Integrated, Structure-Based Approach for Lead-to-Candidate Optimization of Novel CERB Inhibitors that Disrupt Phosphoprotein-Protein Interactions 台大生化分生所-曾秀如
中研院生化所-陳慶士
104.7.1 ~
105.12.31

13

東沙珊瑚礁生物地球化學記錄與環境條件對比研究:重建人類活動在南海的影響
Comparative study between the biogeochemical records in Dongsha coral reef and environmental factors: Reconstructing the anthropogenic impact in the South China Sea
台大地質科學系-任昊佳
中研院地球所-黃國芳
104.7.1 ~
105.12.31

14

逆襲健康資源分配的不平等:社會網絡與地理資訊系統的進擊 台大生傳系-謝雨生
中研院社會所-吳齊殷
104.7.1 ~
105.12.31

 

104年度「臺大與中研院創新性合作計畫」即日起至2月26日止受理申請

 

一、 申請資格:臺大及中研院所屬編制內單位之研究人員或專任教師。
二、 今(104)年度收件日期自即日起至2月26日止,申請資料請以所(中心)為單位,彙整後檢附「申請計畫送件單」,送臺大/中研院聯合辦公室辦理 (地址:10617 臺北市羅斯福路四段一號 台灣大學凝態物理新館R1301)。
三、 注意事項:
(一) 計畫參與單位應包含臺大與中研院。
(二) 應繳資料包括:計畫構想書、臺大/中研院合作意願書、雙方合作經費分攤證明、國內外參與人員CV、其他有助審查之資料等。

審查作業以本計畫構想書進行
  不需另外繳交第二階段計畫書

※ 計畫構想書之「計畫願景」、「執行方式」及「預期效益」,
  如有需要可自行增加行列,至多以2頁為宜

(三) 申請資料請繳交紙本及電子檔案(WORD檔)各1份,電子檔案內之簽名處可附上電子簽名。

四、

相關規定及表單下載請洽本網站連結

 


 

Innovative joint program between National Taiwan University and Academia Sinica

Brief introduction:
1. Upper limit: NT$ 4million/year for each project (In principle, the
support of total amount for each project will be disturbed fifty-fifty
to NTU and AS.)

2. In principle, concurrent acceptance of a grant from any other
organization based on a similar proposal/concepts will be declined..

3. In principle, the period of project is limited to two years. When a
multi-year project is approved under special circumstances, annual
reports are required before issuing next year’s fund.

4. A higher priority will be given to collaboration projects involving
different colleges or departments at NTU.

5. A higher priority will be given to projects with excellence in
innovation and possibility in leading the trend in that research
field.

Materials:
1. proposal
2. A letter of intent from collaborators of NTU and AS.
3. A certificate for expenses apportioned between NTU and AS.
4. A CV and publication list is required for all participants.