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1.
In many electroporation applications mass transport in biological tissue is of primary concern. This paper presents a theoretical advancement in the field and gives some examples of model use in electroporation applications. The study focuses on post-treatment solute diffusion.  相似文献   
2.
朱砂叶螨羧酸脂酶最优测试条件的选择   总被引:1,自引:0,他引:1  
应用二次回归通用旋转组合设计,对朱砂叶螨离体羧酸酯酶测试过程中所需缓冲液pH值、恒温时间、反应温度及底物浓度,设立4因子5水平试验,在考虑4个因子主效应和互作效应的情况下,筛选测试羧酸酯酶的最优条件组合。  相似文献   
3.
In patients with dialysis‐related amyloidosis, β2‐microglobulin (β2‐m) is a major structural component of amyloid fibrils. It has been suggested that the partial unfolding of β2‐m is a prerequisite to the formation of amyloid fibrils, and that the folding intermediate trapped by the non‐native trans‐Pro32 isomer leads to the formation of amyloid fibrils. Although clarifying the structure of this refolding intermediate by high resolution NMR spectroscopy is important, this has been made difficult by the limited lifetime of the intermediate. Here, we studied the structure of the refolding intermediate using a combination of amino acid selective labeling with wheat germ cell‐free protein synthesis and NMR techniques. The HSQC spectra of β2‐ms labeled selectively at either phenylalanine, leucine, or valine enabled us to monitor the structures of the refolding intermediate. The results suggested that the refolding intermediate has an overall fold and cores similar to the native structure, but contains disordered structures around Pro32. The fluctuation of the β‐sheet regions especially the last half of the βB strand and the first half of the βE strand, both suggested to be important for amyloidogenicity, may transform β2‐m into an amyloidogenic structure.  相似文献   
4.
Kinetic, thermodynamic, and structural properties of the aminoglycoside N3‐acetyltransferase‐VIa (AAC‐VIa) are determined. Among the aminoglycoside N3‐acetyltransferases, AAC‐VIa has one of the most limited substrate profiles. Kinetic studies showed that only five aminoglycosides are substrates for this enzyme with a range of fourfold difference in kcat values. Larger differences in KM (~40‐fold) resulted in ~30‐fold variation in kcat/KM. Binding of aminoglycosides to AAC‐VIa was enthalpically favored and entropically disfavored with a net result of favorable Gibbs energy (ΔG < 0). A net deprotonation of the enzyme, ligand, or both accompanied the formation of binary and ternary complexes. This is opposite of what was observed with several other aminoglycoside N3‐acetyltransferases, where ligand binding causes more protonation. The change in heat capacity (ΔCp) was different in H2O and D2O for the binary enzyme–sisomicin complex but remained the same in both solvents for the ternary enzyme–CoASH–sisomicin complex. Unlike, most other aminoglycoside‐modifying enzymes, the values of ΔCp were within the expected range of protein‐carbohydrate interactions. Solution behavior of AAC‐VIa was also different from the more promiscuous aminoglycoside N3‐acetyltransferases and showed a monomer‐dimer equilibrium as detected by analytical ultracentrifugation (AUC). Binding of ligands shifted the enzyme to monomeric state. Data also showed that polar interactions were the most dominant factor in dimer formation. Overall, thermodynamics of ligand‐protein interactions and differences in protein behavior in solution provide few clues on the limited substrate profile of this enzyme despite its >55% sequence similarity to the highly promiscuous aminoglycoside N3‐acetyltransferase. Proteins 2017; 85:1258–1265. © 2017 Wiley Periodicals, Inc.  相似文献   
5.
The Orai1 Ca2+ permeable ion channel is an important component of store operated Ca2+ entry (SOCE) in cells. It’s over-expression in basal molecular subtype breast cancers has been linked with poor prognosis, making it a potential target for drug development. We pharmacologically characterised a number of reported inhibitors of SOCE in MDA-MB-231 breast cancer cells using a convenient Fluorescence Imaging Plate Reader (FLIPR) assay, and show that the rank order of their potencies in this assay is the same as those reported in a wide range of published assays. The assay was also used in a screening project seeking novel inhibitors. Following a broad literature survey of classes of calcium channel inhibitors we used simplified ligand structures to query the ZINC on-line database, and following two iterations of refinement selected a novel Orai1-selective dichlorophenyltriazole hit compound. Analogues of this were synthesized and evaluated in the FLIPR assay to develop structure–activity relationships (SAR) for the three domains of the hit; triazole (head), dichlorophenyl (body) and substituted phenyl (tail). For this series, the results suggested the need for a lipophilic tail domain and an out-of-plane twist between the body and tail domains.  相似文献   
6.
Summary The inheritance of freezing resistance in interspecific F1 hybrid families of Eucalyptus encompassing 27 different species combinations and a range of levels of hardiness was examined. Freezing resistance was assessed by determining the temperatures required to cause either 30% (T30), 40% (T40), or 50% (T50) leakage of electrolytes from excised leaf discs subjected to artificial freezing. Highly significant variation in freezing resistance occurred between species; the maximum difference between parents in any specific combination was over 9°C (E. gunnii x E. globulus). Freezing resistance was inherited in a predominantly additive manner in interspecific hybrids, although there was a tendency towards partial dominance toward the more sensitive species in some combinations (e.g., E. nitens x E. Globulus, E. nitens x E. camaldulensis, E. gunnii x E. globulus). The full expression of this genetic variation appeared to increase with hardiness and in some cases appeared to vary with ontogeny. Estimates of individual narrow-sense heritability of freezing resistance for pure E. nitens families were h 2 = 0.66±0.44 and 0.46±0.44. Across all species combinations examined, the heritability of F1 family means estimated from midparent regression was h 2 = 0.76±0.06 and h 2 = 0.89±0.06 for T40 and T50 values, respectively. The advantage of using selected parents for interspecific hybridization is demonstrated and the implications of these results for breeding for freezing resistance in Eucalyptus are discussed.  相似文献   
7.
Summary The freezing tolerance of cabbage petioles and asparagus shoot apexes was increased by preincubation with 0.8 M sugar solutions. In cabbage petioles with an initial freezing tolerance of –3 °C (temperature for 50% cell survival), as determined by both electrolyte leakage and fluorescein diacetate vital staining, the freezing tolerance was increased to –13 °C by incubation with sorbitol solutions for 3 h. In meristematic cells of asparagus shoot apexes with an initial freezing tolerance of –7.5 °C, as determined by fluorescein diacetate vital staining, the freezing tolerance was increased to –30 °C by incubation with 0.8 M sugar solutions for 3 h, although other cells in the shoot apexes were killed by higher freezing temperatures. During incubation of both cabbage petioles and asparagus shoot apexes with sugar solutions, sugars were intracellularly taken up by osmotically induced fluid-phase endocytotic vesicles, as indicated by comovement of Lucifer Yellows carbohydrazide (LYCH) observed with a confocal laser scanning microscope. The amounts of intracellularly taken up sugars increased concomitantly with the formation of endocytotic vesicles depending on the time of incubation in parallel with a gradual increase of freezing tolerance. However, the endocytotic vesicles and their contents were retained not only after prolonged incubation after maximum freezing tolerance had been achieved but also after recovery of these tissue cells to isotonic conditions or after freeze-thawing. These results suggest that although sugars are intracellularly taken up by endocytotic vesicles, they might be sequestered within vesicles, casting doubt on their protective role to the plasma membranes as a main site of freezing injury. The pretreatment with 1 mMp-chloromercuribenzenesulfonic acid (PCMBS), an inhibitor of sugar transport, reduced the amounts of intracellular sugar uptake without affecting the formation of endocytotic vesicles, suggesting that sugars were, at least partly, taken up by sugar transporters. In the pretreatment with PCMBS, the freezing tolerance of incubated tissues with sugar solutions was significantly reduced, although addition of PCMBS per se did not affect survival. These results suggest that sugars taken up by sugar transporters, rather than sugars taken up by endocytotic vesicles, are mainly responsible for the increased freezing tolerance of cabbage petioles and asparagus shoot apexes. Furthermore, we aimed to study the occurrence of fluid-phase endocytosis with LYCH in an isotonic condition. Our results indicated that uptake of LYCH by fluid-phase endocytotic vesicles was not detected microscopically in isotonic condition, although LYCH was spectrofluorimetrically taken up in isotonic condition. Spectrofluorimetric uptake of LYCH was inhibited by addition of probenecid, an anion transport inhibitor. These results suggest that in cabbage petioles and asparagus shoot apexes, LYCH is taken up by anion transport but not by fluid-phase endocytosis in isotonic condition, and uptake of LYCH by fluid-phase endocytosis is restricted to occur only in hypertonic condition.Abbreviations CLSM confocal laser scanning microscope - FDA fluorescein diacetate - LYCH Lucifer Yellow carbohydrazide - PCMSB p-chloromercuribenzenesulfonic acid - TEL50 temperature at which 50% electrolyte leakage occurred  相似文献   
8.
The structural characterization of small integral membrane proteins pose a significant challenge for structural biology because of the multitude of molecular interactions between the protein and its heterogeneous environment. Here, the three‐dimensional backbone structure of Rv1761c from Mycobacterium tuberculosis has been characterized using solution NMR spectroscopy and dodecylphosphocholine (DPC) micelles as a membrane mimetic environment. This 127 residue single transmembrane helix protein has a significant (10 kDa) C‐terminal extramembranous domain. Five hundred and ninety distance, backbone dihedral, and orientational restraints were employed resulting in a 1.16 Å rmsd backbone structure with a transmembrane domain defined at 0.40 Å. The structure determination approach utilized residual dipolar coupling orientation data from partially aligned samples, long‐range paramagnetic relaxation enhancement derived distances, and dihedral restraints from chemical shift indices to determine the global fold. This structural model of Rv1761c displays some influences by the membrane mimetic illustrating that the structure of these membrane proteins is dictated by a combination of the amino acid sequence and the protein's environment. These results demonstrate both the efficacy of the structural approach and the necessity to consider the biophysical properties of membrane mimetics when interpreting structural data of integral membrane proteins and, in particular, small integral membrane proteins.  相似文献   
9.
本文对等温自由生长和强制性溶液生长的等电溶菌酶的晶体形态进行了研究,发现这些形态变化与溶液相的流动密切相关,指出生物晶体生长停止是由于生长晶体周围的溶质贫乏造成的;通过某些手段减薄或消除这一溶质贫乏区,就可以保证晶体的持续生长。本文的研究对改善大尺寸晶体的生长提供了一条途径。  相似文献   
10.
1IntreductionTheliteratUreonmulti-Criteriondecisionmaking(MCDM)problemshas~tremendouslyintherecentpast.TwomajorareashaveevolvedwhiChbothconcentrateondecisionmakingwithseveralcriteria:multiobjectivedecisionmaking(MODM)andmulti-attributedecisionmaking(MADM).TheformerconcentratesoncontinuousdecisionspaceandthelatterfocusesonproblemswithdiscreteSPace.FuzzysettheoryhascontributedtoMODMproblemsaswellastheMADMProblems.ThegeneralMODMproblemcanbedeft.edLllasfollows:Twostagescangenerallybe…  相似文献   
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