共查询到20条相似文献,搜索用时 15 毫秒
1.
Borelli C Ruge E Lee JH Schaller M Vogelsang A Monod M Korting HC Huber R Maskos K 《Proteins》2008,72(4):1308-1319
Proteolytic activity is an important virulence factor for Candida albicans (C. albicans). It is attributed to the family of the secreted aspartic proteinases (Saps) from C. albicans with a minimum of 10 members. Saps show controlled expression and regulation for the individual stages of the infection process. Distinct isoenzymes can be responsible for adherence and tissue damage of local infections, while others cause systemic diseases. Earlier, only the structures of Sap2 and Sap3 were known. In our research, we have now succeeded in solving the X-ray crystal structures of the apoenzyme of Sap1 and Sap5 in complex with pepstatin A at 2.05 and 2.5 A resolution, respectively. With the structure of Sap1, we have completed the set of structures of isoenzyme subgroup Sap1-3. Of subgroup Sap4-6, the structure of the enzyme Sap5 is the first structure that has been described up to now. This facilitates comparison of structural details as well as inhibitor binding modes among the different subgroup members. Structural analysis reveals a highly conserved overall secondary structure of Sap1-3 and Sap5. However, Sap5 clearly differs from Sap1-3 by its electrostatic overall charge as well as through structural conformation of its entrance to the active site cleft. Design of inhibitors specific for Sap5 should concentrate on the S4 and S3 pockets, which significantly differ from Sap1-3 in size and electrostatic charge. Both Sap1 and Sap5 seem to play a major part in superficial Candida infections. Determination of the isoenzymes' structures can contribute to the development of new Sap-specific inhibitors for the treatment of superficial infections with a structure-based drug design program. 相似文献
2.
贵州喀斯特石漠化地区植物多样性与土壤理化性质 总被引:26,自引:0,他引:26
以贵州典型喀斯特石漠化生态系统环境为研究对象,运用野外取样调查和实验室检测分析方法,研究不同等级石漠化环境植物多样性和土壤理化性质特征及其相关性;运用空间替代时间方法,探讨石漠化演替过程中植物多样性和土壤理化性质的响应,旨在为贵州乃至整个中国西南喀斯特森林生态保护和石漠化生态系统恢复重建提供理论支撑。结果表明:1)石漠化环境植物群路组成简单,物种丰富度也很低,且随着石漠化程度增加,植被物种组成呈递减趋势;不同等级石漠化环境植物多样性具有显著差异,均匀度指数变化与石漠化等级演替明显耦合,显示了随石漠化程度增加而减小的变化趋势。2)不同等级石漠化环境土壤理化性质存在显著差异,随着石漠化程度增加,土壤理化性质显示了先退化后改善的响应过程。土壤有机质、氮素、毛管持水量、容重和孔隙度与植物多样性具有明显的相关性,在改善土壤理化性质和促进植物多样性恢复方面起着关键作用。3)主成分分析表明,土壤有机质、氮素、钾素、持水状况、孔隙度和植物多样性均匀度指数等是基于土壤理化性质和植物多样性评价石漠化程度的关键指标。基于上述结果,进一步阐述了石漠化演替过程中植物多样性和土壤理化性质的变化规律和响应机制。研究结果对我国西南喀斯特森林生态保护和石漠化生态系统恢复重建具有一定的理论意义和实践指导价值。 相似文献
3.
Shapvalova KB 《Rossi?skii fiziologicheski? zhurnal imeni I.M. Sechenova / Rossi?skaia akademiia nauk》2004,90(12):1485-1499
The study shows that, in spite of high criteria of performing inctrumental reflex, the ability to repeat the reflex performance, the increase in tonic component of the response, a dostinct diagonal pattern of posture readjustment, and the local (not diffuse) projection of the mass centre position upon tensoplatforms of anterior paws were only observed after a prolonged training that led to automation of the skill. The instrumental response automation effect could be obtained at once following a bilateral microinjection of carbacholine into the neostriatum. The same albeit a weaker effect could be obtained with bilateral microinjections of D2 Dopamine receptor blocking agent Raclopride into the neostriatum. Bilateral injections of Pyrenzepine yield an opposite result: an increase in the physical component of the response < a disorder in the diagonal pattern of posture readjustment, and a diffuse nature of projection of the mass centre position of anterior paws on tensoplatforms. Indirect efferent output of the neostriatum seems to play an important role in motor instrumental reflex as well as in the process of automation of the motor skill. 相似文献
4.
Main and interaction effects of acetic acid, furfural, and p-hydroxybenzoic acid on growth and ethanol productivity of yeasts 总被引:5,自引:0,他引:5
The influence of the factors acetic acid, furfural, and p-hydroxybenzoic acid on the ethanol yield (YEtOH) of Saccharomyces cerevisiae, bakers' yeast, S. cerevisiae ATCC 96581, and Candida shehatae NJ 23 was investigated using a 2(3)-full factorial design with 3 centrepoints. The results indicated that acetic acid inhibited the fermentation by C. shehatae NJ 23 markedly more than by bakers' yeast, whereas no significant difference in tolerance towards the compounds was detected between the S. cerevisiae strains. Furfural (2 g L-1) and the lignin derived compound p-hydroxybenzoic acid (2 g L-1) did not affect any of the yeasts at the cell mass concentration used. The results indicated that the linear model was not adequate to describe the experimental data (the p-values of curvatures were 0.048 for NJ 23 and 0.091 for bakers' yeast). Based on the results from the 2(3)-full factorial experiment, an extended experiment was designed based on a central composite design to investigate the influence of the factors on the specific growth rate (mu), biomass yield (Yx), volumetric ethanol productivity (QEtOH), and YEtOH. Bakers' yeast was chosen in the extended experiment due to its better tolerance towards acetic acid, which makes it a more interesting organism for use in industrial fermentations of lignocellulosic hydrolysates. The inoculum size was reduced in the extended experiment to reduce any increase in inhibitor tolerance that might be due to a large cell inoculum. By dividing the experiment in blocks containing fermentations performed with the same inoculum preparation on the same day, much of the anticipated systematic variation between the experiments was separated from the experimental error. The results of the fitted model can be summarised as follows: mu was decreased by furfural (0-3 g L-1). Furfural and acetic acid (0-10 g L-1) also interacted negatively on mu. Furfural concentrations up to 2 g L-1 stimulated Yx in the absence of acetic acid whereas higher concentrations decreased Yx. The two compounds interacted negatively on Yx and YEtOH. Acetic acid concentrations up to 9 g L-1 stimulated QEtOH, whereas furfural (0-3 g L-1) decreased QEtOH. Acetic acid in concentrations up to 10 g L-1 stimulated YEtOH in the absence of furfural, and furfural (0-2 g L-1) slightly increased YEtOH in the absence of acetic acid whereas higher concentrations caused inhibition. Acetic acid and furfural interacted negatively on YEtOH. 相似文献
5.
FROZEN THIN SECTIONS OF FRESH TISSUE FOR ELECTRON MICROSCOPY, WITH A DESCRIPTION OF PANCREAS AND LIVER 总被引:13,自引:7,他引:6
下载免费PDF全文
![点击此处可从《The Journal of cell biology》网站下载免费的PDF全文](/ch/ext_images/free.gif)
A. Kent Christensen 《The Journal of cell biology》1971,51(3):772-804
A simple method has been developed that allows frozen thin sections of fresh-frozen tissue to be cut on a virtually unmodified ultramicrotome kept at room temperature. A bowl-shaped Dewar flask with a knifeholder in its depths replaces the stage of the microtome; a bar extends down into the bowl from the microtome's cutting arm and bears the frozen tissue near its lower end. When the microtome is operated, the tissue passes a glass or diamond knife in the depths of the bowl as in normal cutting. The cutting temperature is maintained by flushing the bowl with cold nitrogen gas, and can be set anywhere from about -160°C up to about -30°C. The microtome is set for a cutting thickness of 540–1000 A. Sections are picked up from the dry knife edge, and are placed on membrane-coated grids, flattened with the polished end of a copper rod, and either dried in nitrogen gas or freeze-dried. Throughout the entire process the tissue is kept cold and does not come in contact with any solvent. The morphology seen in frozen thin sections of rat pancreas and liver generally resembles that in conventional preparations, although freezing damage and low contrast limit the detail that can be discerned. Among unusual findings is a frequent abundance of mitochondrial granules in material prepared by this method. 相似文献
6.
It is becoming increasingly clear that there is crosstalk between the apoptotic and autophagic pathways, with autophagy helping to contribute to cell death by providing energy to allow the energy-requiring programmed cell death process to complete, as well as degrading cellular material in its own right. Recent evidence has suggested that Atg proteins can themselves be targets of caspases, providing potential regulation of autophagy as well as uncovering novel functions for fragments derived from Atg proteins. However, to date there has not been a detailed examination of which Atg proteins may be the targets of which death proteases. We show that the majority of human Atg (hAtg) proteins can be cleaved by calpain 1, which is activated in some apoptotic paradigms, as well as other forms of death. We also show that hAtg3 is cleaved by caspases-3, -6 and -8, hAtg6 (Beclin 1) is cleaved by caspase-3 and -6, while hAtg9, hAtg7 and the hAtg4 homologues can be cleaved by caspase-3. Cleavage of Beclin 1 was also seen in apoptosis of HeLa cells induced by staurosporine and TRAIL, along with cleavage of Atg3 and Atg4C. There were subtle effects of caspase inhibition on GFP-LC3 lipidation but more marked effects on the formation of GFP-LC3 puncta (a marker of autophagosome formation) and p62 degradation, indicating that caspase cleavage of autophagy-related proteins can affect the autophagic process. Notably we show that p62 is a target for caspase-6 and -8 cleavage. 相似文献
7.
《Autophagy》2013,9(8):1042-1056
It is becoming increasingly clear that there is crosstalk between the apoptotic and autophagic pathways, with autophagy helping to contribute to cell death by providing energy to allow the energy-requiring programmed cell death process to complete, as well as degrading cellular material in its own right. Recent evidence has suggested that Atg proteins can themselves be targets of caspases, providing potential regulation of autophagy as well as uncovering novel functions for fragments derived from Atg proteins. However, to date there has not been a detailed examination of which Atg proteins may be the targets of which death proteases. We show that the majority of human Atg (hAtg) proteins can be cleaved by calpain 1, which is activated in some apoptotic paradigms, as well as other forms of death. We also show that hAtg3 is cleaved by caspases-3, -6 and -8, hAtg6 (Beclin 1) is cleaved by caspase-3 and -6, while hAtg9, hAtg7 and the hAtg4 homologues can be cleaved by caspase-3. Cleavage of Beclin 1 was also seen in apoptosis of HeLa cells induced by staurosporine and TRAIL, along with cleavage of Atg3 and Atg4C. There were subtle effects of caspase inhibition on GFP-LC3 lipidation but more marked effects on the formation of GFP-LC3 puncta (a marker of autophagosome formation) and p62 degradation, indicating that caspase cleavage of autophagy-related proteins can affect the autophagic process. Notably we show that p62 is a target for caspase-6 and -8 cleavage. 相似文献
8.
The speed of the human genome project (Lander, E. S., Linton, L. M., Birren, B., Nusbaum, C. et al., Nature 2001, 409, 860-921) was made possible, in part, by developments in automation of sequencing technologies. Before these technologies, sequencing was a laborious, expensive, and personnel-intensive task. Similarly, automation and robotics are changing the field of proteomics today. Proteomics is defined as the effort to understand and characterize proteins in the categories of structure, function and interaction (Englbrecht, C. C., Facius, A., Comb. Chem. High Throughput Screen. 2005, 8, 705-715). As such, this field nicely lends itself to automation technologies since these methods often require large economies of scale in order to achieve cost and time-saving benefits. This article describes some of the technologies and methods being applied in proteomics in order to facilitate automation within the field as well as in linking proteomics-based information with other related research areas. 相似文献
9.
Occurrence, metabolism, metabolic role, and industrial uses of bacterial polyhydroxyalkanoates. 总被引:76,自引:0,他引:76
下载免费PDF全文
![点击此处可从《Microbiological reviews》网站下载免费的PDF全文](/ch/ext_images/free.gif)
Polyhydroxyalkanoates (PHAs), of which polyhydroxybutyrate (PHB) is the most abundant, are bacterial carbon and energy reserve materials of widespread occurrence. They are composed of 3-hydroxyacid monomer units and exist as a small number of cytoplasmic granules per cell. The properties of the C4 homopolymer PHB as a biodegradable thermoplastic first attracted industrial attention more than 20 years ago. Copolymers of C4 (3-hydroxybutyrate [3HB]) and C5 (3-hydroxyvalerate [3HV]) monomer units have modified physical properties; e.g., the plastic is less brittle than PHB, whereas PHAs containing C8 to C12 monomers behave as elastomers. This family of materials is the centre of considerable commercial interest, and 3HB-co-3HV copolymers have been marketed by ICI plc as Biopol. The known polymers exist as 2(1) helices with the fiber repeat decreasing from 0.596 nm for PHB to about 0.45 nm for C8 to C10 polymers. Novel copolymers with a backbone of 3HB and 4HB have been obtained. The native granules contain noncrystalline polymer, and water may possibly act as a plasticizer. Although the biosynthesis and regulation of PHB are generally well understood, the corresponding information for the synthesis of long-side-chain PHAs from alkanes, alcohols, and organic acids is still incomplete. The precise mechanisms of action of the polymerizing and depolymerizing enzymes also remain to be established. The structural genes for the three key enzymes of PHB synthesis from acetyl coenzyme A in Alcaligenes eutrophus have been cloned, sequenced, and expressed in Escherichia coli. Polymer molecular weights appear to be species specific. The factors influencing the commercial choice of organism, substrate, and isolation process are discussed. The physiological functions of PHB as a reserve material and in symbiotic nitrogen fixation and its presence in bacterial plasma membranes and putative role in transformability and calcium signaling are also considered. 相似文献
10.
R Powers D S Garrett C J March E A Frieden A M Gronenborn G M Clore 《Biochemistry》1992,31(17):4334-4346
The assignment of the 1H, 15N, 13CO, and 13C resonances of recombinant human interleukin-4 (IL-4), a protein of 133 residues and molecular mass of 15.4 kDa, is presented based on a series of 11 three-dimensional (3D) double- and triple-resonance heteronuclear NMR experiments. These studies employ uniformly labeled 15N- and 15N/13C-labeled IL-4 with an isotope incorporation of greater than 95% for the protein expressed in yeast. Five independent sequential connectivity pathways via one-, two-, and three-bond heteronuclear J couplings are exploited to obtain unambiguous sequential assignments. Specifically, CO(i)-N(i + 1),NH(i + 1) correlations are observed in the HNCO experiment, the C alpha H(i), C alpha (i)-N(i + 1) correlations in the HCA(CO)N experiment, the C alpha(i)-N(i + 1),NH(i + 1) correlations in the HNCA and HN(CO)CA experiments, the C alpha H(i)-N(i + 1),NH(i + 1) correlations in the H(CA)NH and HN(CO)HB experiments, and the C beta H(i)-N(i + 1),NH(i + 1) correlations in the HN(CO)HB experiments. The backbone intraresidue C alpha H(i)-15N(i)-NH(i) correlations are provided by the 15N-edited Hartmann-Hahn (HOHAHA) and H(CA)NH experiments, the C beta H(i)-15N(i)-NH(i) correlations by the 15N-edited HOHAHA and HNHB experiments, the 13C alpha(i)-15N(i)-NH(i) correlations by the HNCA experiment, and the C alpha H(i)-13C alpha(i)-13CO(i) correlations by the HCACO experiment. Aliphatic side-chain spin systems are assigned by 3D 1H-13C-13C-1H correlated (HCCH-COSY) and total correlated (HCCH-TOCSY) spectroscopy. Because of the high resolution afforded by these experiments, as well as the availability of multiple sequential connectivity pathways, ambiguities associated with the limited chemical shift dispersion associated with helical proteins are readily resolved. Further, in the majority of cases (88%), four or more sequential correlations are observed between successive residues. Consequently, the interpretation of these experiments readily lends itself to semiautomated analysis which significantly simplifies and speeds up the assignment process. The assignments presented in this paper provide the essential basis for studies aimed at determining the high-resolution three-dimensional structure of IL-4 in solution. 相似文献
11.
WHO Collaborating Centres (CCs) form part of an international collaborative network set up by WHO in support of its mandated programme at the country, intercountry, regional, interregional and global levels, as appropriate. As part of its mandate in the area of biologicals, WHO has broadened the scope of its work and has expanded the range of activities devoted to the establishment of international standards for vaccines. In line with global immunization goals, the need for standards for evaluation of quality, safety and efficacy of new vaccines, as well as those that have been in use for a long time, has significantly increased. Furthermore, complex issues related to new production methodologies, more sophisticated techniques for characterization and laboratory testing, and for nonclinical and clinical evaluation of vaccines have raised a number of regulatory challenges for WHO when requested to assist its Member States. In this context, CCs in the area of standardization of vaccines and biotherapeutics (excluding blood products) have provided technical assistance and have broadened the scope of their work over time. In the area of standardization and regulatory evaluation of vaccines, WHO currently has six CCs as well as one candidate centre for which the designation process has been initiated and a further three candidate centres with great potential. The purpose of the meeting held on 24–26 April 2012 was to improve understanding of WHO's priorities in setting standards, to facilitate their implementation, and to increase transparency of the roles and responsibilities of CCs. The meeting was also an excellent opportunity to explore possibilities for improving collaboration between WHO and CCs, as well as among CCs themselves by working as a CC network. All CCs expressed a wish for increased interaction, information-sharing, collaboration and other ways of working together that may lead to cross-fertilization between the CCs. Synergy was recognized as a significant mechanism for leveraging existing resources in responding to global public health challenges and in addressing WHO's priorities. Agreement was reached for operating as a network of CCs. 相似文献
12.
Herman van Langen Tessa Castelijn 《Physica medica : PM : an international journal devoted to the applications of physics to medicine and biology : official journal of the Italian Association of Biomedical Physics (AIFB)》2009,25(4):207-211
In many X-ray clinics, the traditional photographic film has been replaced by an imaging plate (IP). The IP is re-usable and the purpose of this study was to test if image deterioration occurred after successive uses of the IP. The emphasis is placed on the efficiency of image formation and on image uniformity.In a cross-sectional study, 21 clinically used IPs were exposed with a standardized phantom imaging protocol. These IPs were in clinical use between one month and two years and the IPs were exposed between 191 and 3787 times. After digitizing, the mean pixel value (MPV) in a predefined image area was determined. The relation between MPV and IP uses was assessed.In a second experiment, image uniformity of 30 other clinically used IPs was visually inspected for artifacts on a diagnostic monitor. These IPs were in clinical use between one week and two years and exposed between 76 and 5373 times.The first experiment showed that no significant deterioration of the MPV with increasing usage count of the IP was present (p = 0.15). The second experiment showed the appearance of clinically relevant artifacts on the IP before 3000 uses.It was concluded that the efficiency of the image formation process does not significantly deteriorate after successive use of IPs and is therefore not expected to limit their life span. Mechanical handling in the digitizer of the used system seems to set a limit to IP durability. Uniformity should therefore be checked regularly in clinical quality control. 相似文献
13.
Li Liang-Qian 《植物分类学报:英文版》1988,26(3):189-204
The geographical distribution of Aconitum in the Sino-Himalayan subregion is
analysed in the present paper on the basis of taxonomy and relationship between the infrageneric taxa. Asaresult, some conclusions may be arrived as follows:
1. The Sino-Himalayan subregion is the frequency centre and the diversity centre of the
genus. For analysis, the distribution area of the genus are assigned to three floristic regions,
viz. 1) the East-Asian floristic region, consisting of the Sino-Himalayan subregion and SinoJapanese subregion, 2) the Euro-Siberian region and 3) the North-American region (Table 1).
In the East-Asian floristic region, the Sino-Himalayan subregion comprises 3 subgenera, about
5 sections, about 13 series and nearly 180 species. However, the Sino-Japanese subregion has
only 2 subgenera, 2 sections. 6 series and about 50 species. The Euro-Siberian region has 2
subgenera, 2 sections, about 9 series and nearly 70 species. The North-American region has
2 subgenera (one of the 2 subgenera has only 1 species), 1 section, 1-2 series and about 26
species. Obviously, the Sino-Himalayan subregion is the richest in taxa.
2. The Sino-Himalayan subregion is not only the preservation centre of the primitive
groups and species, but also an actively differentiating region. Largely in the Sino-Himalayan
subregion occurs primitive or more primitive tava in the genus, such as Sect. Fletcherum, Sect.
Alatosperum and Sect. Sinaconitum, Ser. Tangutica and Ser. Brunnea etc.: A fletcherianum,
A. novoluridum, A. chrysotricum, A. brevicalcaratum, A. polycarpus, A. nagarum, A. tanguti cum, A. hookeri, A. naviculare, A. violaceum, etc. On the other hand, the Sino-Himalayan
subregion also has the most or relatively advanced taxa, represented by the annual monotypic
subgenus Gymnaconitum, A. spiripetalum, A. hamatipetalum and A. bulbitiferum ect. About
50 infraspecific taxa occur in the subregion. For example, A. hemsleyanum has 8 varities, A.
franchetii has varities and A. nagarum 1 varity and 2 forms, ect.
3. In the region under discussion the genus Aconitum shows remarkable endemism.
The endemic taxa include 3 sections (Sect. Fletcherum, Sect. Alatosperum and Sect. Sina conitum), 3 series (Ser. Brevicalcarata, Ser. Crassiflora and Ser. Bullatifolia) and nearly 150
species, among which primitive and advanced ones are both present.
4. The pattern of geographical distribution of the genus Aconitum shows remarkable
relationship between latitude and altitude. The majority of species of this genus prefer habi tats with a cool and more or less constantly moist climate. In the Sino-Japanese subregion, with
a higher latitude, the genus has an altitude range of 500-1500 m, whereas in the Sino-Hima-layan subregion the range is 2900-5000 m.
To sum up, the Sino-Himalayan subregion is the diversity centre, the frequency centre, the
differentiation centre, the preservation centre of the primitive taxa and the centre of endemism
of the genus Aconitum, and its development in this subregion has probably been accelerated
by the lift of the Himalayas and the complicated environmental conditions. 相似文献
14.
Temperature regulation of microtiter plates for enzyme assays 总被引:1,自引:0,他引:1
To facilitate the use of microtiter plates as vessels for enzyme assays, an incubator has been designed to maintain the wells of the microtiter plates at the appropriate temperature. The temperature variation within a single well was +/- 0.02 degrees (standard error of the mean) at 25 degrees C and +/- 0.02 degrees at 37 degrees C. The temperature variation was the same for internal and peripheral wells within the plates, although the internal wells were approximately 0.14 degrees C warmer than the outer wells at 25 degrees C and 0.68 degrees C cooler at 37 degrees C. The overall uncertainty (95% confidence interval) of the well temperature in a typical plate was +/- 0.4 degrees C at 25 degrees C and +/- 0.7 degrees C at 37 degrees C. This uncertainty is realistic for routine enzyme determinations, as opposed to precise studies. The incubator was designed to allow access to the plates from above so that additions could be made during the incubation. To demonstrate the suitability of microtiter plates and the incubator for enzyme determinations, this method was used to measure the activity of myeloperoxidase and alpha-naphthylbutyryl esterase. 相似文献
15.
Hematospermia in the dog usually occurs secondary to benign prostatic hypertrophy or trauma of the penis or prepuce during semen collection. Regarding the difficulty of removing blood cells from a hematospermic sample, the present study was performed to determine whether blood contaminated ejaculates can still be chilled (4 degrees C) or frozen (-196 degrees C) without an additional decrease in sperm quality. In the first experiment, blood additions of up to 10% exerted no negative effects on the functional characteristics of canine spermatozoa cooled (4 degrees C) and stored for 4 days in an egg-yolk-Tris extender. In contrast, in experiment 2, blood admixtures of 4% or more clearly caused negative effects on cryopreserved (-196 degrees C) spermatozoa, mainly on the motility parameters, on the membrane integrity and on the acrosomal status of the spermatozoa. In experiment 3, we showed that these negative effects of blood admixture on cryopreserved spermatozoa were mainly associated with the red blood cells (RBCs) whereas the addition of plasma, serum or inactivated serum exerted little or no negative effect. Moreover, in experiment 4, we showed that 58.3+/-11.6% of the RBCs hemolysed after a freeze-thaw process. In experiment 5, a clear and negative effect of hemoglobin on cryopreserved canine spermatozoa was observed. We conclude that the presence of up to 10% blood is not detrimental for the storage of chilled canine spermatozoa and that the detrimental effects of blood on cryopreserved spermatozoa are at least partly attributable to the high amount of hemoglobin originating from the RBC hemolysis observed after freezing and thawing. 相似文献
16.
Labeling DNA with stable isotopes to measure cell proliferation can be a technique as effective as 3H-thymidine labeling without the limitations imposed by using radioisotopes. Here, we investigated the relative efficiency of four nonradioactive precursors to DNA: [1-13C]-glycine, [1,2-13C2]-glycine, [U-13C]-glucose, and [U-13C, 15N]-thymidine. The efficiency of incorporation for each of these labeled precursors in HEP G2 cells in culture has been studied. When considering the actual costs of in vivo experiments in which large doses of labeled material are needed, economical constraints may play an important role in defining a practical method. Therefore, the economics of this process were also considered. Using the enrichment per dollar for whichever nucleoside had the highest incorporation in a given experiment, glycine is about five times more economical as a label than thymidine and eight times more economical than glucose in these cells. 相似文献
17.
MOTIVATION: AutoGrid/AutoDock is one of the most popular software packages for docking, but its automation is not trivial for tasks such as (1) the virtual screening of a library of ligands against a set of possible receptors; (2) the use of receptor flexibility and (3) making a blind-docking experiment with the whole receptor surface. This is an obstacle for research teams in the fields of Chemistry and the Life Sciences who are interested in conducting this kind of experiment but do not have enough programming skills. To overcome these limitations, we have designed BDT, an easy-to-use graphic interface for AutoGrid/AutoDock. AVAILABILITY: BDT is available for free, upon request, for non-commercial research. 相似文献
18.
A modification of an established procedure for autoradiographic processing of cultured cells is described. This method eliminates the need for pipetting each individual well and also for cutting and dismantling the multiwell plate for slide preparation. In this procedure the entire plate can be processed as a single unit and the cells can be analyzed in situ, thus eliminating the time consuming pipetting and cutting procedures. Furthermore, the entire experiment can be filed without use of additional slides or storage boxes. Hence, this is a simpler, time conserving, and economical way to process large numbers of cultures for thymidine labeling indices. 相似文献
19.
Small proteins provide convenient models for computational studies of protein folding and stability, which are usually compared with experimental data. Until recently, the unfolding of Trp-cage was considered to be a two-state process. However, no direct experimental evidence for this has been presented, and in some cases, the contrary has been suggested. To elucidate a detailed unfolding mechanism, we studied the thermodynamics of unfolding of Trp-cage by differential scanning calorimetry (DSC) and circular dichroism (CD) spectroscopy. The observation that at low temperatures only approximately 90-95% of Trp-cage exists in the native conformation presented an analytical challenge. Nevertheless, it was found that the DSC and CD data can be fitted simultaneously to the same set of thermodynamic parameters. The major uncertainty in such a global fit is the heat capacity change upon unfolding, DeltaCp. This can be circumvented by obtaining DeltaCp directly from the difference between heat capacity functions of the native and unfolded states. Using such an analysis it is shown that Trp-cage unfolding can be represented by a two-state model with the following thermodynamic parameters: Tm = 43.9 +/- 0.8 degrees C, DeltaH(Tm) = 56 +/- 2 kJ/mol, DeltaCp = 0.3 +/- 0.1 kJ/(mol.K). Using these thermodynamic parameters it is estimated that Trp-cage is marginally stable at 25 degrees C, DeltaG(25 degrees C) = 3.2 +/- 0.2 kJ/mol, which is only 30% more than the thermal fluctuation energy at this temperature. 相似文献
20.
Wang B Miskolizie M Kotovych G Pulay P 《Journal of biomolecular structure & dynamics》2002,20(1):71-80
Calculated and experimental (1)H, (13)C and (19)F chemical shifts were compared in BKM-824, a cyclic bradykinin antagonist mimic, c[Ava(1)-Igl(2)-Ser(3)-DF5F(4)-Oic(5)-Arg(6)] (Ava=5-aminovaleric acid, Igl=alpha-(2-indanyl)glycine, DF5F=pentafluorophenylalanine, Oic=(2S,3aS,7aS)-octahydroindole-2-carboxylic acid). The conformation of BKM-824 has been studied earlier by NMR spectroscopy (M. Miskolzie et al., J. Biomolec. Struct. Dyn. 17, 947-955 (2000)). All NMR structures have qualitatively the same backbone structure but there is considerable variation in the side chain conformations. We have carried out quantum mechanical optimization for three representative NMR structures at the B3LYP/6-31G* level, constraining the backbone dihedral angles at their NMR structure values, followed by NMR chemical shift calculations at the optimized structures with the 6-311G** basis set. There is an intramolecular hydrogen bond at Ser(3) in the optimized structures. The experimental (13)C chemical shifts at five C(alpha) positions as well as at the Cbeta, Cgamma and Cdelta position of Ava(1), which forms part of the backbone, are well reproduced by the calculations, confirming the NMR backbone structure. A comparison between the calculated and experimental H(beta) chemical shifts in Igl(2) shows that the dominant conformation at this residue is gauche. Changes of proton chemical shifts with the scan of the chi(1) angle in DF5F(4) suggest that chi(1)180 degrees. The calculated (1)H and (13)C chemical shifts are in good agreement with experiment at the rigid residue Oic(5). None of the models gives accurate results for Arg(6), presumably because of its positive charge. Our study indicates that calculated NMR shifts can be used as additional constraints in conjunction with NMR data to determine protein conformations. However, to be computationally effective, a database of chemical shifts in small peptide fragments should be precalculated. 相似文献