• BMB Reports
• /
• 제39권5호
• /
• pp.530-536
• /
• 2006

Thermal conformational changes of human serum albumin (HSA) in phosphate buffer, 10 mM at pH = 7 are investigated using differential scanning calorimetric (DSC), circular dichroism (CD) and UV spectroscopic methods. The results indicate that temperature increment from $25^{\circ}C$ to $55^{\circ}C$ induces reversible conformational changes in the structure of HSA. Conformational change of HSA are shown to be a three-step process. Interestingly, melting temperature of the last domain is equal to the maximum value of fever in pathological conditions, i.e. $42^{\circ}C$. These conformational alterations are accompanied by a mild alteration of secondary structures. Study of HSA-SDS (sodium dodecyl sulphate) interaction at $45^{\circ}C$ and $35^{\circ}C$ reveals that SDS affects the HSA structure at least in three steps: the first two steps result in more stabilization and compactness of HSA structure, while the last one induces the unfolding of HSA. Since HSA has a more affinity for SDS at $45^{\circ}C$ compared to $35^{\circ}C$, It is suggested that the net negative charge of HSA is decreased in fever, which results in the decrease of HSA-associated cations and plasma osmolarity, and consequently, heat removal via the increase in urine volume.

• 한국방사선학회논문지
• /
• 제7권6호
• /
• pp.433-439
• /
• 2013

본 연구에서는 집속형 초음파를 이용하는 뇌 질환 치료의 연구를 위한 인체 두부 유사 팬텀을 개발하였다. 문헌 연구를 통하여 피부 조직, 두개골, 뇌 조직의 음향학적 및 물리적 특성을 조사하였으며 근사한 값을 가지는 적합한 각 조직 대체 물질을 제시하였다. 피부 조직의 경우 글리세롤 기반 연부 조직 유사 팬텀의 성분비를 조정하여 실제 조직과 유사한 음향학적 특성을 가지도록 하였으며 고분자 합성수지의 음향학적 특성을 측정하여 두개골 유사 물질로써의 적합성을 평가하였다. 뇌 조직은 투명한 egg white 팬텀을 이용하여 집속형 초음파의 가열 특성을 확인할 수 있도록 하였다. 또한 뇌 질환 치료 프로토콜 개발을 위한 시험 조사가 가능하도록 대체 물질들을 결합한 두부 유사 팬텀을 제작하였고 제작된 팬텀의 유효성 및 활용성 평가를 위해 초음파 조사 조건에 따른 팬텀의 변성을 관찰하였다.

• Journal of the Korean Wood Science and Technology
• /
• 제29권3호
• /
• pp.112-122
• /
• 2001

Laccase enzymes from Cerrena unicolor and Trametes versicolor were immobilized on the activated glass beads (CPG), silica gel (SG) and soil (SL). The heterogeneous matrices were activated by ${\gamma}$-aminopropyltriethoxysilane (APTES) and glutaraldehyde (GA), and their surfaces were coated by keratin (KER) on activated or non-activated CPG, SG and SL. The laccase activities were tested in the aqueous solution for the native and immobilized preparations using different pH and temperature conditions. By keratin coating on supports, in the cases of CPG-KER and SL-KER, the immobilization yield was increased from about 80% to 90%. Moreover, much less protein was immobilized in keratin coated matrices than in inorganic ones alone (e.g. on CPG-KER 57.6%, whereas on CPG alone 80.6%). Laccase immobilization on keratin coated inorganic matrices was generally more effective than that of non-coated matrices. Concerned to pH dependency, the optima pH for immobilized laccases generally shifted towards to higher values, 5.5-5.8 and even 5.9 in the case of keratin for C. unicolor and from 5.3 to 5.7 for T. versicolor, respectively, and decreased less gradually both in acidic and alkaline regions. The immobilized laccase was more stable against thermal denaturation. This seems particularly true at $75^{\circ}C$ in the case of C. unicolor, where the activity of immobilized enzyme is > 50% higher than that of the free enzyme. For T. versicolor the respective values were $65^{\circ}C$, and 50%.

• 생명과학회지
• /
• 제14권2호
• /
• pp.309-314
• /
• 2004

cAMP수용성 단백질(CRP)은 E. coli의 100가지 이상의 유전자 전자조절에 관계된다. CRP는 dimer로 존재하며 cAMP의 결합으로 활성인 형태로 전환된다. 이중체인 CRP 단백질의 열 안정성과 구조 전이의 연구에 효과적인 온도 기울기 전기영동장치를 이용하여 확인하였다. 본 연구에서 야생형과 S83C CRP 단백질의 melting temperature (Tm)는 산성인 완충용액[89.8 mM Glycine, 24mM Boric acid (pH 5.8)]에서 57$\pm$1(야생형 CRP)과 55$\pm$1$^{\circ}C$ (S83G CRP)였다. 그리고 온도에 따른 CRP 단백질의 구조변화도 protease digestion과 CD spectropolarimeter을 이용하여 확인하였다.

• Fisheries and Aquatic Sciences
• /
• 제12권1호
• /
• pp.6-15
• /
• 2009

Biochemical and functional properties of acid-soluble collagen (ASC) and pepsin-soluble collagen (PSC) from rockfish skin were characterized. Yield of PSC (90.0%) was higher than that of ASC (63.2%). Both ASC and the PSC consisted of ${\alpha}1$ and ${\alpha}2$ chains, and $\alpha$-cross-linked components. According to the results of hydroxylation of proline and lysine, and FT-IR, no difference between the helical structure of ASC and PSC was identified. Thermal denaturation temperature (TDT) of ASC from rockfish skin was $22.8^{\circ}C$, the same as exhibited in PSC. Both ASC and PSC were higher in water absorption capacity (WAC) and oil absorption capacity (OAC) than other vegetable proteins. According to the results of emulsifying activity (EA) and cooking stability (CS), both ASC and PSC from rockfish skin were inferior compared to the commercial emulsifier (Tween-80). The results of FT-IR suggested that the structure of PSC was slightly different when compared to that of ASC. No differences in solubility were established between ASC and PSC from rockfish skin at various pH and NaCl concentrations.

• BMB Reports
• /
• 제33권6호
• /
• pp.493-498
• /
• 2000

Aquifex pyrophilus, a hyperthermophilic bacterium, has a serine-type protease that is located at the cell wall fraction with a mature size of 43 kDa. Molecular cloning of the protease gene revealed that it has an ORF of 619 amino acids with homologous catalytic site of serine-type proteases [Choi, I.-G., Bang, W.-K., Kim, S.-H., Yu, G. Y., J. Biol. Chem. (1999), Vol. 274, pp. 881-888]. Constructs containing different regions of the protease gene, including a alanine-substituted mutant at the active site serine, were constructed, and the factors affecting the expression level of the cloned protease gene in E. coli were examined. The presence of the C-terminus hydrophobic region of the protease hindered over-expression in E. coli. Also, the proteolytic activity of the expressed protein appeared to toxic to E. coli. An inactive form that deleted both of the N-terminal signal sequence and the C-terminal polar residues was over-expressed in a soluble form, purified to homogeneity, and its thermostability examined. The purified protein showed three disulfide bonds and three free sulfhydryl group. The thermal denaturation temperature of the protein was measured around $90^{\circ}C$ using a differential scanning calorimeter and circular dichroism spectrometry. The disulfide bonds were hardly reduced in the presence of reducing agents, suggesting that these disulfide bonds were located inside of the protein surface.

• Molecules and Cells
• /
• 제38권2호
• /
• pp.187-194
• /
• 2015

Thioredoxin (TRX) is a disulfide reductase present ubiquitously in all taxa and plays an important role as a regulator of cellular redox state. Recently, a redox-independent, chaperone function has also been reported for some thioredoxins. We previously identified nodulin-35, the subunit of soybean uricase, as an interacting target of a cytosolic soybean thioredoxin, GmTRX. Here we report the further characterization of the interaction, which turns out to be independent of the disulfide reductase function and results in the co-localization of GmTRX and nodulin-35 in peroxisomes, suggesting a possible function of GmTRX in peroxisomes. In addition, the chaperone function of GmTRX was demonstrated in in vitro molecular chaperone activity assays including the thermal denaturation assay and malate dehydrogenase aggregation assay. Our results demonstrate that the target of GmTRX is not only confined to the nodulin-35, but many other peroxisomal proteins, including catalase (AtCAT), transthyretin-like protein 1 (AtTTL1), and acyl-coenzyme A oxidase 4 (AtACX4), also interact with the GmTRX. Together with an increased uricase activity of nodulin-35 and reduced ROS accumulation observed in the presence of GmTRX in our results, especially under heat shock and oxidative stress conditions, it appears that GmTRX represents a novel thioredoxin that is co-localized to the peroxisomes, possibly providing functional integrity to peroxisomal proteins.

• 대한전기학회:학술대회논문집
• /
• 대한전기학회 2006년도 제37회 하계학술대회 논문집 C
• /
• pp.1629-1630
• /
• 2006

This paper describes a micro total analysis system ($\mu$ TAS) for detecting and digesting the target protein which includes a bead based temperature controllable microchip and computer based controllers for temperature and valve actuation. We firstly combined the temperature control function with a bead based microchip and realized the on-chip sequential reactions using two kinds of beads. The PEG-grafted bead, on which RNA aptamer was immobilized, was used for capturing and releasing the target protein. The target protein can be chosen by the type of RNA aptamer. In this paper, we used the RNA aptamer of HCV replicase. The trypsin coated bead was used for digesting the released protein prior to the matrix assisted laser desorption ionization time of flight mass spectrometer (MALDI TOF MS). Heat is applied for release of the captured protein binding on the bead, thermal denaturation and trypsin digestion. PDMS microchannel and PDMS micro pneumatic valves were also combined for the small volume liquid handling. The entire procedures for the detection and the digestion of the target protein were successfully carried out on a microchip without any other chemical treatment or off-chip handling using $20\;{\mu}l$ protein mixture within 20 min. We could acquire six matched peaks (7% sequence coverage) of HCV replicase.

• Journal of Microbiology and Biotechnology
• /
• 제30권12호
• /
• pp.1905-1911
• /
• 2020

Homoserine dehydrogenase (HSD) catalyzes the reversible conversion of ʟ-aspartate-4-semialdehyde to ʟ-homoserine in the aspartate pathway for the biosynthesis of lysine, methionine, threonine, and isoleucine. HSD has attracted great attention for medical and industrial purposes due to its recognized application in the development of pesticides and is being utilized in the large scale production of ʟ-lysine. In this study, HSD from Bacillus subtilis (BsHSD) was overexpressed in Escherichia coli and purified to homogeneity for biochemical characterization. We examined the enzymatic activity of BsHSD for ʟ-homoserine oxidation and found that BsHSD exclusively prefers NADP+ to NAD+ and that its activity was maximal at pH 9.0 and in the presence of 0.4 M NaCl. By kinetic analysis, Km values for ʟ-homoserine and NADP+ were found to be 35.08 ± 2.91 mM and 0.39 ± 0.05 mM, respectively, and the Vmax values were 2.72 ± 0.06 μmol/min-1 mg-1 and 2.79 ± 0.11 μmol/min-1 mg-1, respectively. The apparent molecular mass determined with size-exclusion chromatography indicated that BsHSD forms a tetramer, in contrast to the previously reported dimeric HSDs from other organisms. This novel oligomeric assembly can be attributed to the additional C-terminal ACT domain of BsHSD. Thermal denaturation monitoring by circular dichroism spectroscopy was used to determine its melting temperature, which was 54.8℃. The molecular and biochemical features of BsHSD revealed in this study may lay the foundation for future studies on amino acid metabolism and its application for industrial and medical purposes.

• 한국축산식품학회지
• /
• 제44권5호
• /
• pp.1028-1039
• /
• 2024

Alternative protein sources with greater nutritional value and a lower environmental footprint have recently attracted interest in the production of meat substitutes. However, it is required that these alternatives mimic the texture and structure of meat. This study investigated varying ratios of textured vegetable proteins (TVP) to Tenebrio molitor larvae (brown mealworm; TM) with the addition of transglutaminase (TG) to determine the quality characteristics of these emulsions. The results demonstrated low protein solubility of the emulsions as TVP content increased. Furthermore, when the proportion of TM was high, the TG-treated emulsion had a low pH. Additionally, when there was a high TM ratio to TVP in the TG treatment, the emulsions demonstrated better thermal stability and water holding capacity. Regarding the rheological properties of the emulsion, both the frequency-dependent storage modulus (G') and loss modulus (G'') increased as the proportion of TVP in the emulsion increased with and without the addition of TG. Differential scanning calorimetry analyses demonstrated two protein denaturation peaks in all treatments, with high peak temperatures for both treatments with a high proportion of TM. The hardness and chewiness of the emulsion were highest in the treatment (T6 and T8) with TG, and the gumminess of the emulsion was greatest when TM only or when equal ratios of TVP and TM were treated with TG, respectively. In conclusion, the addition of TM to TVP with TG improves the overall texture of the protein mixture, making it a suitable meat alternative.