• 제목/요약/키워드: solid and protein

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고체상 핵자기공명 분광법을 이용한 막단백질의 구조연구 (Structural Studies of Membrane Protein by Solid-state NMR Spectroscopy)

  • 김용애
    • 분석과학
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    • 제17권5호
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    • pp.388-392
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    • 2004
  • genomics의 정보해석이나 신경전달물질 또는 약의 전달체계에서 아주 중요한 역할을 담당하는 막단백질의 구조연구는 기존의 X-ray나 용액상 핵자기공명분광법으로 수행하기 어려우나 지방질 이분자층이나 여러분자층에서 움직이지 않게 정렬시킨 단백질시료를 이용하여 특이하게 고안된 home-built solid-state NMR probe를 이용하면 구조를 연구할 수 있다. 이 논문에서는 박테리오파지인 pf1의 growth, 분리, 정제 및 pf1에서의 coat protein의 분리, 정제과정과 최종적으로 분리 정제된 pf1의 coat protein의 인산지방질 이분자층에서의 구조를 고체상 핵자기공명 분광법을 이용하여 연구하고자 한다.

Construction of 1H-15N Double Resonance Solid-State NMR Probe for Membrane Proteins in Aligned Bicelles

  • Park, Tae-Joon;Kim, Ji-Sun;Um, Seung-Hoon;Kim, Yong-Ae
    • Bulletin of the Korean Chemical Society
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    • 제31권5호
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    • pp.1187-1191
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    • 2010
  • $^1H-^{15}N$ heteronuclear dipolar coupling solid-state NMR experiments on lipid bilayer or bicelle samples are very useful for the structural studies of membrane proteins. However, to study these biological samples using solid-state NMR, a specific probe with high efficiency and high capability is required. In this paper, we describe the optimized design, construction, and efficiency of a 400 MHz wide-bore $^1H-^{15}N$ solid-state NMR probe with 5-mm solenoidal rf coil for high power, multi-pulse sequence experiments, such as 2D PISEMA or 2D SAMMY.

Low-ε Static Probe Development for 15N-1H Solid-state NMR Study of Membrane Proteins for an 800 MHz NB Magnet

  • Park, Tae-Joon;Choi, Sung-Sub;Jung, Ji-Ho;Park, Yu-Geun;Kim, Yongae
    • Bulletin of the Korean Chemical Society
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    • 제34권3호
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    • pp.823-826
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    • 2013
  • A low-${\varepsilon}$ solid-state NMR(Nuclear Magnetic Resonance) probe was developed for the spectroscopic analysis of two-dimensional $^{15}N-^1H$ heteronuclear dipolar coupling in dilute membrane proteins oriented in hydrated and dielectrically lossy lipid environments. The system employed a 800 MHz narrow-bore magnet. A solenoid coil strip shield was used to reduce deleterious RF sample heating by minimizing the conservative electric fields generated by the double-tuned resonator at high magnetic fields. The probe's design, construction, and performance in solid-state NMR experiments at high magnetic fields are described here. Such high-resolution solid-state NMR spectroscopic analysis of static oriented samples in hydrated phospholipid bilayers or bicelles could aid the structural analysis of dilute biological membrane proteins.

Salt-Induced Protein Precipitation in Aqueous Solution: Single and Binary Protein Systems

  • Kim, Sang-Gon;Bae, Young-Chan
    • Macromolecular Research
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    • 제11권1호
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    • pp.53-61
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    • 2003
  • A molecular-thermodynamic model is developed for the salt-induced protein precipitation. The protein molecules interact through four intermolecular potentials. An equation of state is derived based on the statistical mechanical perturbation theory with the modified Chiew's equation for the fluid phase, Young's equation for the solid phase as the reference system and a perturbation based on the protein-protein effective two body potential. The equation of state provides an expression for the chemical potential of the protein. In a single protein system, the phase separation is represented by fluid-fluid equilibria. The precipitation behaviors are simulated with the partition coefficient at various salt concentrations and degree of pre-aggregation effect for the protein particles. In a binary protein system, we regard the system as a fluid-solid phase equilibrium. At equilibrium, we compute the reduced osmotic pressure-composition diagram in the diverse protein size difference and salt concentrations.

표고버섯의 고체배양에 의한 단백 다당류 생산 (Production of Protein-bound Polysaccharides by Solid-substrate Fementation of Lentinus edodes)

  • 박경숙
    • 한국식품영양학회지
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    • 제11권6호
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    • pp.667-672
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    • 1998
  • The possibility of solid-substrate fermentation of Lentinus edoes for the productin of protein-boud polysaccharides (PBP) was studied. Zeolite and orchid-pot soil were used as solid materials for the culture because of the desirable physical properties. Sucrose and starch were good carbon sources for the production of PBP by the solid-substrate fermentatin of L. edodes. Among the nitrogen source, bactosoyton was very effective for the PBP production. The optimum pH for solid-substrate fementation for the production of PBP was at pH of 5.5. The PBP production reached to 5∼5.5mg per 100g solid-substrate.

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식용 왕달팽이의 영양성분과 단백질 품질 (Nutrient Composition and Protein Quality of Giant Snail Products)

  • Mi-kyung Lee;Jeung-hye Moon;Hong-Soo Ryu
    • 한국식품영양과학회지
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    • 제23권3호
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    • pp.453-458
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    • 1994
  • 체색이 다른 세종류(백색육, 황색육, 회색육)의 식용 달팽이(Giant snail, Achatine fulica)의 일반성분과 무기질을 정량하여 이의 식품학적 가능성을 예측하였으며, 효소를 이용한 단백소화율(in vitro protein digestibility), 단백효율비(C-PER, computed protein efficiency ratio) 및 typsin inibitor 함량을 측정하여 이의 단백질 품질을 평가하였다. 또한 전처리 및 가공조건을 달리한 달팽이 제품의 단백질 품질을 측정하여 바람직한 처리조건을 찾으려 했으며, 달팽이육 중에 다량 포함되어 있는 점액성 물질이 다른 단백질의 소화율에 미치는 영향도 아울러 실험하였다. 1. 식용왕달팽이의 가식부는 39% 정도로서 해산패류보다 10%정도 높았으며 가식부 100g당 11.5~13.7%의 단백질, 0.9~1.3%정도의 지질이 함유되어 있다. 무기질로서는 칼슘, 칼륨 및 마그네슘이 풍부하였고 특히 칼슘함량은 굴(oyster)에 비교하여 5~10배 높았다. 2. 아미노산의 함량과 그 조성은 백색육, 회색육, 황색육이 모두 비슷하였고, 주요 아미노산은 aspaltic acid, glutamic acid, proline과 glycine으로 총아미노산의 52.5%를 차지하였고 특히 필수아미노산 함량이 총아미노산의 45~46%를 차지하여 균형잡힌 단백질로 판별되었다. 3. 생달팽이육의 단백질 소화율은 76.6%(회색육)~81%(황색육과 백색육)였으며 단백효율비는 2.24(백색육)~2.36(황색육) 범위였으나, trypsin inhibitor함량은 체색에 따라 다양하였다(백색윤 22.7mg/g solid, 회색육 28.97mg/g solid 및 황색육 36.75mg/g solid). 4. 10% 식염수에 5분간 침지한 후 10분간 boiling한 달팽이육의 소화율은 생육에 비해 1.7%(백색육)~5.7%(회색육)로 증가하였고, trypsin inhibitor 함량은 12.62mg/g solid (백색육)과 16.40mg/g solid (회색육)로 크게 감소하였으며, C-PER은 가공조건에 따라 변동이 없었다. 5. 생달팽이육에 다량 함유된 점액성물질은 여러가지 단백질의 소화율 2% (ANRC casein)~11%(filefish protein)로 감소시켰다.

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Construction of 19F-13C Solid-State NMR Probe for 400MHz Wide-Bore Magnet

  • Jeong, Ji-Ho;Park, Yu-Geun;Choi, Sung-Sub;Kim, Yongae
    • 한국자기공명학회논문지
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    • 제17권2호
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    • pp.81-85
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    • 2013
  • Various fluorine-containing materials are used in electronic devices like LCD display panels and Li-ion batteries. The structural conformation of fluorine in fluorinated materials is an important contributing factor that influences the chemical and physical properties. The conformation can be changed by heat and stress during manufacture or use. Understanding the conformational changes is critical for understanding the performance and durability of electronic devices. Solid-state NMR spectroscopy could be widely used for the analysis of various fluorine-containing materials for electronic devices. However, conventional CPMAS probes cannot be used for in-situ analysis of fluorine-containing electronic devices like LCD panels and Li-ion batteries. In this paper, we show the design, construction, and optimization of a $^{19}F-^{13}C$ double-resonance solid-state NMR probe for a 400MHz wide-bore magnet with a flat square coil for in-situ analysis of fluorine-containing electronic devices without observing fluorine background signals. This custom-built probe does not show any fluorine background signals, and can have higher efficiency for lossy samples.

Variable Temperature High-Resolution 19F MAS Solid-State NMR Characterization of Fluorocarbon Rubbers

  • Park, Tae-Joon;Choi, Sung-Sub;Kim, Ji-Sun;Kim, Yong-Ae
    • Bulletin of the Korean Chemical Society
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    • 제32권7호
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    • pp.2345-2350
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    • 2011
  • Variable temperature high-resolution $^{19}F$ magic angle spinning (MAS) solid-state NMR spectroscopy was used to characterize fluorocarbon (FKM) rubbers. The high-resolution spectra of copolymers made from two monomers, vinylidene fluoride and hexafluoropropene, and terpolymers composed of vinylidene fluoride, hexafluoropropene, and tetrafluoroethylene, were obtained using MAS speeds of up to 18 kHz combined with high temperatures of up to 200 $^{\circ}C$ at a magnetic field strength of 9.4 Tesla. From these high resolution solid-state NMR spectra, we were able to assign the spectral peaks and differentiate the copolymer FKM from the terpolymer FKM. We also determined quantitatively the monomer compositions of each FKM rubber.

Solid-phase refolding of poly-lysine tagged fusion protein of hEGF and angiogenin

  • Park, Sang-Joong;Ryu, Kang;Chai, Young-Gyu;Kweon, Oh-Byung;Park, Seung-Kook;Lee, Eun-Kyu
    • 한국생물공학회:학술대회논문집
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    • 한국생물공학회 2001년도 추계학술발표대회
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    • pp.197-203
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    • 2001
  • A fusion protein, consisting of human epidermal growth factor as a recognition domain and human angiogenin as a toxin domain, can be used as a targeted therapeutic against breast cancer cells among others. The fusion protein was expressed as inclusion body in recombinant E. coli, and when the conventional, solution-phase refolding process was used the refolding yield was very low due to severe aggregation, probably due to the opposite surface charge due to vastly different pI values of each domain. Solid-phase refolding process exploiting ionic interactions between the solid matrix and the protein was tried, but the ionic binding yield was very low regardless of the resins and pH conditions used. To provide higher affinity toward the solid matrix, six lysine residues were tagged to the N -terminus of the hEGF domain When the cation exchange resins such as heparin- or CM-Sepharose were used as the matrix, the adsorption capacity increased 2.5-3 times and the subsequent refolding yield increased nearly IS times compared to the conventional process.

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Solid-phase Refolding of Poly-lysine Tagged Fusion Protein of hEGF and Angiogenin

  • Park Sang Joong;Ryu Kang;Suh Chang Woo;Chai Young Gyu;Kwon Oh Byung;Park Seung Kook;Lee Eun Kyu
    • Biotechnology and Bioprocess Engineering:BBE
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    • 제7권1호
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    • pp.1-5
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    • 2002
  • A fusion protein, consisting of a human epidermal growth factor (hEGF) as the recognition domain and human angiogenin as the toxin domain, can be used as a targeted therapeutic against breast cancer cells among others. The fusion protein was expressed as inclusion body in recombinant E. coli, and when the conventional, solution-phase refolding process was used the refolding yield was very low due to severe aggregation. It was probably because of the opposite electric charge at a neutral pH resulting from the vastly different pI values of each domain. The solid-phase refolding process that exploited the ionic interactions between ionic exchanger surface and the fusion protein was tried, but the adsorption yield was also very low, below $ 30\%$, regardless of the resins and pH conditions used. Therefore, to provide a higher ionic affinity toward the solid matrix, six lysine residues were tagged to the N-terminus of the hEGF domain. When heparin-Sepharose was used as the matrix, the adsorption capacity increased 2.5-3 times to about $88\%$. Besides the intrinsic affinity of angiogenin to heparin, the poly-lysine tag provided additional ionic affinity. And the subsequent refolding yield increased nearly 13-fold, from ca. $4.8\%$ in the conventional refolding of the untagged fusion protein to $63.6\%$. The process was highly reproducible. The refolded protein in the column eluate retained RNase bioactivity of angiogenin.