생체유기분자가 결합된 인지질막을 이용한 다기능성 나노구조의 합성 및 응용

Abstract

본 논문에서는 생체유기분자가 결합된 인지질막을 이용하여 다기능성 나노구조체를 합성하였고, 그 구조 및 특성을 grazing incidence small and wide angle x-ray scattering (GISAXS, GIWAXS), x-ray reflectivity (XRR), atomic force microscopy (AFM), transmission electron microscopy (TEM) 그리고 differential scanning calorimeter (DSC)로 분석하였고, 나노 및 바이오 기술로의 응용가능성을 보았다. 인지질은 다형성(polymorphism)을 갖고 주변환경에 따라 다양한 상으로 존재할 수 있기 때문에, transmembrane 단백질 중 하나로써 구하기 용이하며 막 내에서 안정한 구조를 갖는 gramicidin과 광합성에 필수적인 성분인 chlorophyll의 두 가지 생체유기분자와 결합하여 생체막을 형성하는데 활용되었다. 생체유기분자의 첨가가 dipalmitoyl phosphatidylcholine (DPPC) 인지질 막의 구조에 어떤 영향을 미치는지 알아보기 위해 GISAXS, XRR 그리고 AFM 분석을 시행하였고, 막의 표면 형태, 막 두께와 거칠기, 막의 내부 형태, lamellar 구조의 존재유무 등의 구조적 해석이 가능하였다. 두 생체유기분자 gramicidin과 chlorophyll의 첨가는 막의 lamellar 구조의 파괴 및 표면 형태의 평탄함을 초래하였지만, 막의 두께가 변화하는 경향성은 두 생체유기분자의 크기 차이로 인해 다르게 나타났다. 생체막의 인지질에 대한 상변이 현상은 GIWAXS와 DSC로 관찰되었으며, 생체유기분자의 양이 증가할수록 인지질의 alkyl-chain의 운동이 방해를 받게 되고, 따라서 온도에 따른 상변이가 점차적으로 일어나지 않는다는 결론에 도출하게 되었다. DPPC 인지질막에 gramicidin을 농도별로 첨가하여, 증착되는 금속나노입자와의 상관관계를 알아보았다. Gramicidin이 첨가되지 않은 단단한 gel 상태의 인지질막의 경우, Ag, Sn, Al 그리고 Au 금속 원자들이 쉽게 핵형성을 할 수 없기 때문에, 금속나노입자의 형성 밀도가 낮았으며, gramicidin이 첨가된 인지질막의 경우, 금속 원자들과의 친화도가 증가하여 보다 고밀도의 금속나노입자가 형성되었다. 그리고 나노입자의 크기는 금속의 종류 및 gramicidin의 농도에 따라 3nm에서 15nm로 다양했으며, 나노입자의 증착밀도는 gramicidin의 농도에 크게 영향을 받았다. 이 연구는 상대적으로 생체친화적인 막에 여러 금속 및 금속산화 나노입자들을 합성할 수 있는 쉽고 편리한 방법을 제공하였다. 인지질막과 transmembrane 단백질을 결합시켜서, 물에 녹지 않는 생체친화적 전극을 제작하였다. Gramicidin을 첨가함으로써 solid-supported 인지질막의 소수성기를 크게 증대시킬 수 있었고, 이렇게 소수성이 증가된 인지질막은 물속 뿐만 아니라 공기/물 계면에서 안정적으로 구조를 유지하였으며, 이는 바이오센서, 효소연료전지, surface enhanced raman scattering (SERS) 기판 등으로의 응용가능성을 보여주었다. 앞선 연구들을 바탕으로, 생체유기분자가 결합된 인지질막에 나노미터 크기의 입자들을 형성시키고, 인지질 분자들로 encapsulation된 나노입자를 합성하였다. Solid-supported 생체막은 생체유기분자와 인지질이 포함된 용액을 실리콘 기판 위에 스핀코팅함으로써 형성하였고, 여기에 물리 증착법 (physical vapor deposition)을 이용하여 금속나노입자를 형성하였다. 인지질 분자가 금속입자 주위에 자발적으로 encapsulation되면서 나노입자의 추가적인 성장을 막고 그 크기 및 모양을 특정하게 결정하였고, 이 인지질이 encapsulation된 나노입자의 형태 및 크기는 온도와 습도를 변화시킴으로써 제어할 수 있었을 뿐만 아니라, 인지질의 종류 및 증착 두께를 조절함으로써도 제어가 가능하였다. 그리고 인지질이 encapsulation된 금속나노입자를 용매에 재분산하였고, Langmuir-Blodgett 방법을 도입하여 대면적 그리고 고밀도의 자가정렬된 나노입자 필름을 형성하였다. 자가정렬된 나노입자 필름을 형성하는 이 방법은 단순분산된 필름을 얻는 기존 방법에 비해 경제적이고 편리할 뿐만 아니라 효과적이며, 이렇게 형성된 대면적, 고밀도의 필름은 광필터, 광센서, SERS 기판 및 chlorophyll 분자가 결합된 광화학적 에너지 재배 시스템으로의 응용 가능성을 보여주었다. 위에서 언급된 실험결과들을 통해서 생체유기분자의 첨가가 인지질막의 구조와 성질을 변화시킬 수 있다는 걸 확인하였고, 이렇게 제어된 생체막은 다기능성 장치의 나노구조 및 추가적인 연구를 통한 나노바이오 융합기술로의 응용이 가능할 것이다. |In this dissertation, multi-functional nanostructures were synthesized using phospholipid membrane modified with bioorganic molecules and their structures and properties were characterized by grazing incidence small and wide angle x-ray scattering (GISAXS, GIWAXS), x-ray reflectivity (XRR), atomic force microscopy (AFM), Transmission electron microscopy (TEM) and differential scanning calorimeter (DSC) for applications toward nano- and bio-technology. Since phospholipid molecules are highly polymorphic so that they can be formed in a variety of ways, giving rise to different phases, phospholipid was chosen to produce biomembranes modified by two different bioorganic molecules: i.e. gramicidin which is one of affordable and stable transmembrane proteins and chlorophyll which is one of essential pigments in the photosynthesis. To understand how bioorganic molecule addition affects the structure of planar dipalmitoyl phosphatidylcholine (DPPC) multilayer phospholipid membrane, GISAXS, XRR and AFM were introduced. These tools enable determination of the structural details of the bioorganic molecule-modified biomembrane which included the surface morphology of the membrane, the membrane thickness and roughness, the texture of the membrane, and the presence of lamellar structure. Both gramicidin and chlorophyll additions resulted in the perturbation of the lamellar structure in the membrane and the smoothed surface morphology. However, the tendency of the thickness change was different because of the size difference between gramicidin and chlorophyll molecules. The phase transition of the phospholipids in the biomembrane was studied by GIWAXS and DSC. As the amount of bioorganic molecule addition was increased, the main phase transition peak of the phospholipids in the biomembrane became faint and shifted to low temperature due to the perturbed alkyl-chain confinement of the phospholipids by the incorporation of the bioorganic molecules. DPPC multilayer phospholipid membrane was structurally modified by introducing a transmembrane protein, gramicidin (up to 25 mol%) to study its effect on the metal nanoparticles deposited on the membrane. Without gramicidin, when 3-nm-thick Ag, Sn, Al and Au were deposited, the nanoparticles hardly nucleated on the DPPC membrane in rigid gel state (except for Au); however, the gramicidin addition dramatically enhanced the DPPC membrane surface’s affinity for metal atoms so that a dense array of metal (Ag, Sn, and Au) or metal-oxide (Al-oxide) nanoparticles was produced on the membrane surface. The particle size ranged from 3 nm to 15 nm depending on the metal and gramicidin concentration whereas the particle density was strongly dictated by the gramicidin concentration. The proposed method provides convenient, generally applicable synthesis route for preparing different metal or metal-oxide nanoparticles on a relatively robust biocompatible membrane. A water-insoluble, biocompatible electrode composed of phospholipid and transmembrane protein was fabricated. The hydrophobicity of solid-supported phospholipid multilayer can be greatly increased by incorporating a transmembrane protein, gramicidin into the lipid membrane. The increased hydrophobicity of the gramicidin-modified lipid membrane allowed the membrane to remain stable at the air/water interface as well as underwater. The enhanced underwater stability of the lipid multilayer substantially broadens the potential application of the lipid multilayer which included biosensing, enzymatic fuel cell and SERS substrate. Based on the preceding research, a novel method of using a solid-supported phospholipid membrane modified with bioorganic molecules as a template to synthesize nanometer-sized particles as well as to make the encapsulation of the resulting particles with phospholipid molecules was proposed. Solid-supported biomembrane was synthesized by spin-coating of phospholipid solution mixed with bioorganic molecules on Si substrate and then metal nanoparticles were deposited on it by physical vapor deposition method. Spontaneous encapsulation of metal particles by the phospholipid molecules would limit the further growth of metal particles beyond an equilibrium size by confining the volume and shape of the metal particles. The shape and size of lipid-encapsulated nanoparticles was controlled not only by temperature and humidity changes but also by a variety of lipids and the change of deposition thickness. Lipid-encapsulated metal nanoparticles were re-dispersed and introduced on Langmuir-Blodgett trough to synthesis a large-area and a high-density self-assembled nanoparticle film on the substrate. This approach to synthesis self-assembled nanoparticle film is economical, effective and simple compared to the conventional methods to produce monodispersed nanoparticles. The proposed large-scale and high-density films can be applied for light filter, light sensor, SERS substrate as well as a photochemical energy harvesting system using chlorophyll molecules. Abovementioned experimental results demonstrated that the structure and the property of phospholipid membrane could be tuned by addition of bioorganic molecules and bioorganic molecule modified biomembrane is a useful tool to enable potential applications of the nanostructure in multi-functional devices. With further research, the proposed applications can serve as a bridge for converging recent developments in nano- and bio-technology.; In this dissertation, multi-functional nanostructures were synthesized using phospholipid membrane modified with bioorganic molecules and their structures and properties were characterized by grazing incidence small and wide angle x-ray scattering (GISAXS, GIWAXS), x-ray reflectivity (XRR), atomic force microscopy (AFM), Transmission electron microscopy (TEM) and differential scanning calorimeter (DSC) for applications toward nano- and bio-technology. Since phospholipid molecules are highly polymorphic so that they can be formed in a variety of ways, giving rise to different phases, phospholipid was chosen to produce biomembranes modified by two different bioorganic molecules: i.e. gramicidin which is one of affordable and stable transmembrane proteins and chlorophyll which is one of essential pigments in the photosynthesis. To understand how bioorganic molecule addition affects the structure of planar dipalmitoyl phosphatidylcholine (DPPC) multilayer phospholipid membrane, GISAXS, XRR and AFM were introduced. These tools enable determination of the structural details of the bioorganic molecule-modified biomembrane which included the surface morphology of the membrane, the membrane thickness and roughness, the texture of the membrane, and the presence of lamellar structure. Both gramicidin and chlorophyll additions resulted in the perturbation of the lamellar structure in the membrane and the smoothed surface morphology. However, the tendency of the thickness change was different because of the size difference between gramicidin and chlorophyll molecules. The phase transition of the phospholipids in the biomembrane was studied by GIWAXS and DSC. As the amount of bioorganic molecule addition was increased, the main phase transition peak of the phospholipids in the biomembrane became faint and shifted to low temperature due to the perturbed alkyl-chain confinement of the phospholipids by the incorporation of the bioorganic molecules. DPPC multilayer phospholipid membrane was structurally modified by introducing a transmembrane protein, gramicidin (up to 25 mol%) to study its effect on the metal nanoparticles deposited on the membrane. Without gramicidin, when 3-nm-thick Ag, Sn, Al and Au were deposited, the nanoparticles hardly nucleated on the DPPC membrane in rigid gel state (except for Au)