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1.
Purification of proteins is commonly a multiple-step process involving size exclusion, ion exchange, affinity, hydrophobic, and other modes of chromatography. In an effort to circumvent the laborious process of collecting the solutes from each column and reintroducing them onto a second column, a valving system is described that directs the samples eluted from a high-performance liquid chromatographic column through a detector with a high-pressure cell into either a second column or into storage loops of a multiloop value. This multiloop value is referred to as a high-pressure fraction collector. After development of the first column is complete, a second solvent can be directed to the second column or high-pressure fraction collector to elute the solutes back through the detector and onto any other column in the system. The process of eluting a sample from a column through a single detector and directing it to the high-pressure fraction collector or any other column in the system may be repeated a number of times. Such valving systems make it possible to chromatograph a single protein component on two or three columns in a short time. 相似文献
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The ultrastructure and systematics of two red colored freshwater cryptomonads, Storeatula rhinosa , sp. nov. and Pyrenomonas ovalis , sp. nov., are described for the first time. Storeatula, which had been described from marine waters only, has a single inner periplast sheet and a fibrous surface periplast component. Cells lack a furrow but possess a gullet, a bilobed chloroplast connected by a pyrenoid and a nucleomorph located in an indentation of the pyrenoid. This freshwater Storeatula possesses the same general features as the marine species, but it has a contractile vacuole and lacks the lobed chloroplast of S. major. P. ovalis has the generic characteristics described for marine species of Rhodomonas. These characteristics include a short furrow, a deep gullet, square inner periplast plates with beveled corners, a slightly fibrillar surface periplast component, a single chloroplast with two lobes connected by a pyrenoidal bridge and a nucleomorph located in an indentation of the pyrenoid. 相似文献
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The ribosomal protein S27a (RPS27a) is cleaved from the fusion protein ubiquitin–RPS27a (Ub–RPS27a). Generally, Ub and RPS27a are coexpressed as a fusion protein but function independently after Ub is cleaved from RPS27a by a deubiquitinating enzyme. As an RP, RPS27a assembles into ribosomes, but it also functions independently of ribosomes. RPS27a is involved in the development and poor prognosis of various cancers, such as colorectal cancer, liver cancer, chronic myeloid leukemia, and renal carcinoma, and is associated with poor prognosis. Notably, the murine double minute 2/P53 axis is a major pathway through which RPS27a regulates cancer development. Moreover, RPS27a maintains sperm motility, regulates winged aphid indirect flight muscle degeneration, and facilitates plant growth. Additionally, RPS27a is a metalloprotein and mercury (Hg) biomarker. In the present review, we described the origin, structure, and biological functions of RPS27a. 相似文献
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由石家庄中国人民解放军白求恩国际和平医院皮肤科李成龙大夫提供的一株人体皮肤病菌经我们研究鉴定为毛霉目(Mucorales)毛霉科(Mucoraceae)根毛霉属(Rhizomucor)的一个种,并定名为多变根毛霉新种(Rhizomucor variabilis Zheng & G.-q.Chen sp. nov.)。据李大夫介绍,这株菌是从一名居住在江苏农村到该医院看病的女病人的手上病部分离的。这个病人没有一般真菌病病人所患有的其他疾病如糖尿病、白血病等等。她也没有患有其他毛霉病,因此她的皮肤毛霉病是原发性的而不是继发性的。我们查阅文献结果,国内由根毛霉属真菌引致的毛霉病过去仅有过一次肺部感染的报道;国外则有过较多次数的由根毛霉引致的人体毛霉病,主要为肺部疾病并可引致继发性的皮肤病,尚未见有由根毛霉属引起的原发性皮肤毛霉病的报道。无论国内、外引起人体毛霉病的根毛霉均为微小根毛霉[Rhizomucor pusillus(Lindt) Schipper,包括Mucor pusillus Lindt,Mucor parasiticus Lucet & Costanin等异名]一种。因此,本病例为我国第二例由根毛霉弓l起的人体毛霉病及第一例由根毛霉引起的人体皮肤毛霉病,同时又是全世界第一例由根毛霉属除微小根毛霉以外的另外一个种引起的人体毛霉病,很可能还是全世界第一例由根毛霉引起的人体原发性皮肤毛霉病。多变根毛霉与根毛霉属内所有过去已报道过的种都有显著差异。它的最适生长温度为24-30℃,最低9℃,最高38℃;其他种均为高温真菌,它们的最高生长温度可达55℃或更高。形态方面,多变根毛霉也与属内其他已知种明显不同。它的菌落高达4-8 mm并呈鲜明的浅黄色;其他已知种菌落低矮,除奈尼塔尔根毛霉(Rhizomucor nainitalensis Joshi)外全部为深暗灰色,奈尼塔尔根毛霉菌落色泽虽然较浅,但为浅灰或灰黄色,与多变根毛霉的鲜明黄色不同。它的假根异常发达并可从菌体的各个部位如菌丝、匍匐丝、孢子枝、孢子囊、囊轴上长出;其他已知种的假根一般都不发达且从未见有从孢子枝、孢子囊、囊轴等处长出的描述。它的孢子枝的分枝常常长于主枝;其他已知种则分枝长度一般不超过主枝。它的囊轴形状多变:球形、近球形、扁球形、卵形、椭圆形、梨形等等,两边对称或不对称,溢缩或不绕缩,纵向深裂或不作纵向深裂;其他已知种的囊轴形状通常为倒卵形至梨形的规则形状。它的囊领明显;其他已知种的囊领均很不明显至缺如。它的孢囊孢子形状和大小变化都较大,卵形、椭圆形、矩圆形、近球形、近三角形或其他各种不规则形状,长度范围2.5-16.5 μm;其他已知种除上面已经提到过的奈尼塔尔根毛霉外,它们的孢囊孢子形状仅限于卵形、椭圆形、近球形等较规则的形状,长度范围总是在3-6 μm范围内,奈尼塔尔根毛霉的孢囊孢子形状虽然多变,但其决度亦在3-6 μm的范围内。此外,多变根毛霉的孢子囊、囊轴、孢囊孢子等各种构造均较大;其他已知种则较小,其中肿梗根毛霉[Rhizomucor tauricus (Milko & Schkurenko) Schipper]虽亦较大,但除此之外与多变根毛霉迥异。多变根毛霉未见有接合孢子,将我们保存的全部微小根毛霉菌株与它分别配对时,或将我们的几对别的属的(+)(-)测试菌株与它分别配对时,均未见形成接合孢子或有任何反应。 相似文献
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Wnt proteins and Wnt signalings have been implicated in a variety of development and cell processes, while aberrant activation of Wnt signaling is linked to a range of cancers in many tissues. In this study, we used the HEK293 cell line to investigate the effects of Wnt3a and Wnt5a on proliferation and apoptosis in a serum starvation culture. After Wnt3a and Wnt5a proteins were expressed, they both promoted the proliferation of HEK293 cells under serum starvation. After 48h of serum starvation, both Wnt3a and Wnt5a inhibited serum starvation-induced apoptosis of HEK293 cells and continued up to 96h. We demonstrated that Wnt3a and Wnt5a can promote proliferation of HEK293 cells and inhibit serum starvation-induced apoptosis, which implies that Wnt3a and Wnt5a can maintain the survival of HEK293 cells under stress, and also provide a novel insight into the role of Wnt3a and Wnt5a and their related signalings in carcinogenesis. 相似文献
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Sec1/Munc18-like (SM) proteins functionally interact with SNARE proteins in vesicular fusion. Despite their high sequence conservation, structurally disparate binding modes for SM proteins with syntaxins have been observed. Several SM proteins appear to bind only to a short peptide present at the N terminus of syntaxin, designated the N-peptide, while Munc18a binds to a 'closed' conformation formed by the remaining portion of syntaxin 1a. Here, we show that the syntaxin 16 N-peptide binds to the SM protein Vps45, but the remainder of syntaxin 16 strongly enhances the affinity of the interaction. Likewise, the N-peptide of syntaxin 1a serves as a second binding site in the Munc18a/syntaxin 1a complex. When the syntaxin 1a N-peptide is bound to Munc18a, SNARE complex formation is blocked. Removal of the N-peptide enables binding of syntaxin 1a to its partner SNARE SNAP-25, while still bound to Munc18a. This suggests that Munc18a controls the accessibility of syntaxin 1a to its partners, a role that might be common to all SM proteins. 相似文献
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Bagley DC Paradkar PN Kaplan J Ward DM 《The Journal of biological chemistry》2012,287(30):25577-25588
Mon1a was originally identified as a modifier gene of vesicular traffic, as a mutant Mon1a allele resulted in increased localization of cell surface proteins, whereas reduced levels of Mon1a showed decreased secretory activity. Here we show that Mon1a affects different steps in the secretory pathway including endoplasmic reticulum-to-Golgi traffic. siRNA-dependent reduction of Mon1a levels resulted in a delay in the reformation of the Golgi apparatus after Brefeldin A treatment. Endoglycosidase H treatment of ts045VSVG-GFP confirmed that knockdown of Mon1a delayed endoplasmic reticulum-to-Golgi trafficking. Reductions in Mon1a also resulted in delayed trafficking from Golgi to the plasma membrane. Immunoprecipitation and mass spectrometry analysis showed that Mon1a associates with dynein intermediate chain. Reductions in Mon1a or dynein altered steady state Golgi morphology. Reductions in Mon1a delayed formation of ERGIC-53-positive vesicles, whereas reductions in dynein did not affect vesicle formation. These data provide strong evidence for a role for Mon1a in anterograde trafficking through the secretory apparatus. 相似文献
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Yi Xiao Fang Liu Qinghong Kong Xinting Zhu Haijuan Wang Sanhua Li Nian Jiang Changyan Yu Liu Yun 《Aging cell》2022,21(3)
Metformin, a widely prescribed first‐line drug for the treatment of type II diabetes mellitus, has been shown to extend lifespan and delay the onset of age‐related diseases. The precisely mechanisms by which these effects are realized remain elusive. We find that metformin exposure is restricted to adults, which is sufficient to extend lifespan. However, limiting metformin exposure to the larvae has no significant effect on Caenorhabditis elegans longevity. Here, we show that after metformin treatment, the level of S‐adenosylmethionine (SAM) is reduced in adults but not in the larvae. Potential mechanisms by which reduced SAM might increase lifespan include altering the histone methylation. However, the molecular connections between metformin, SAM limitation, methyltransferases, and healthspan‐associated phenotypes are unclear. Through genetic screening of C. elegans, we find that metformin promotes the healthspan through an H3K4 methyltransferase/demethylase complex to downregulate the targets, including mTOR and S6 kinase. Thus, our studies provide molecular links between meformin, SAM limitation, histone methylation, and healthspan and elucidate the mode action of metformin‐regulated healthspan extension will boost its therapeutic application in the treatment of human aging and age‐related diseases. 相似文献
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Minimal model of self-replicating nanocells: a physically embodied information-free scenario 总被引:1,自引:0,他引:1
Fellermann H Solé RV 《Philosophical transactions of the Royal Society of London. Series B, Biological sciences》2007,362(1486):1803-1811
The building of minimal self-reproducing systems with a physical embodiment (generically called protocells) is a great challenge, with implications for both theory and applied sciences. Although the classical view of a living protocell assumes that it includes information-carrying molecules as an essential ingredient, a dividing cell-like structure can be built from a metabolism-container coupled system only. An example of such a system, modelled with dissipative particle dynamics, is presented here. This article demonstrates how a simple coupling between a precursor molecule and surfactant molecules forming micelles can experience a growth-division cycle in a predictable manner, and analyses the influence of crucial parameters on this replication cycle. Implications of these results for origins of cellular life and living technology are outlined. 相似文献
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Lamon-Fava S Marcovina SM Albers JJ Kennedy H Deluca C White CC Cupples LA McNamara JR Seman LJ Bongard V Schaefer EJ 《Journal of lipid research》2011,52(6):1181-1187
The aim of this study was to assess the independent contributions of plasma levels of lipoprotein(a) (Lp(a)), Lp(a) cholesterol, and of apo(a) isoform size to prospective coronary heart disease (CHD) risk. Plasma Lp(a) and Lp(a) cholesterol levels, and apo(a) isoform size were measured at examination cycle 5 in subjects participating in the Framingham Offspring Study who were free of CHD. After a mean follow-up of 12.3 years, 98 men and 47 women developed new CHD events. In multivariate analysis, the hazard ratio of CHD was approximately two-fold greater in men in the upper tertile of plasma Lp(a) levels, relative to those in the bottom tertile (P < 0.002). The apo(a) isoform size contributed only modestly to the association between Lp(a) and CHD and was not an independent predictor of CHD. In multivariate analysis, Lp(a) cholesterol was not significantly associated with CHD risk in men. In women, no association between Lp(a) and CHD risk was observed. Elevated plasma Lp(a) levels are a significant and independent predictor of CHD risk in men. The assessment of apo(a) isoform size in this cohort does not add significant information about CHD risk. In addition, the cholesterol content in Lp(a) is not a significant predictor of CHD risk. 相似文献
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Primary structure and functional characterization of rat C5a: an anaphylatoxin with unusually high potency. 总被引:2,自引:0,他引:2
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L. Cui D. F. Carney T. E. Hugli 《Protein science : a publication of the Protein Society》1994,3(8):1169-1177
The anaphylatoxin C5a is a pro-inflammatory factor generated from C5 during complement activation. C5a derived from rat C5 exhibits significantly greater potency compared to C5a from other species. Rat C5a was 25-fold more potent than human C5a for eliciting spasmogenic contraction of guinea pig ileum. Proteolytic removal of the C-terminal arginine of C5a (C5adesArg) reduced spasmogenic potency of rat C5a by only 4-fold compared to a 3,000-fold reduction for human C5adesArg. In addition, rat C5adesArg was 50-fold more potent than human C5adesArg in a guinea pig vascular permeability (in vivo) assay and as a chemotactic factor for human neutrophils. C5a and C5adesArg were purified from zymosan-activated rat serum. Rat C5a, like human C5a, is glycosylated but contains 77 amino acid residues instead of the 74 residues of human C5a. Comparison of the primary structures of rat and human C5a indicated differences at 30 positions including an insert of 3 residues (LLH) in the rat molecule between residue positions 3 and 4 in human C5a. Insertion of residues LLH between Gln-3 and Lys-4 in a recombinant human C5a molecule using site-directed mutagenesis failed to enhance potency. Synthetic C-terminal analogues of rat C5a proved to be measurably more potent than the corresponding human C5a analogues (Ember JA et al., 1993, Protein Sci 2(Suppl 1):159 [Abstr]). We conclude that multiple sequence differences in the C-terminal effector portion and/or elsewhere in rat C5a, but not the LLH insert, account for the significant enhancement in potency of rat C5a over C5a from other species. 相似文献
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Musculoskeletal models are made to reflect the capacities of the human body in general, and often a specific subject in particular. It remains challenging to both model the musculoskeletal system and then fit the modelled muscles to a specific human subject. We present a reduced muscle model, a planar musculoskeletal model, and a fitting method that can be used to find a feasible set of active and passive muscle parameters for a specific subject. At a minimum, the fitting method requires inverse dynamics data of the subject, a scalar estimate of the peak activation reached during the movement, and a plausible initial estimate for the strength and flexibility of that subject. While additional data can be used to result in a more accurate fit, this data is not required for the method solve for a feasible fit. The minimal input requirements of the proposed fitting method make it well suited for subjects who cannot undergo a maximum voluntary contraction trial, or for whom recording electromyographic data is not possible. To evaluate the model and fitting method we adjust the musculoskeletal model so that it can perform an experimentally recorded stoop-lift of a 15 kg box. 相似文献
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Maruyama T Shinohara T Hosogi T Ichinose H Kamiya N Goto M 《Analytical biochemistry》2006,354(1):8-14
Guanine quenching of a fluorescence-labeled DNA probe is a powerful tool for detecting a mutation in a targeted site of a DNA strand. However, a different guanine adjacent to a targeted site can interfere with detection of a point mutation, resulting in unsatisfactory sensitivity. In the current study, we developed a simple method to improve sensitivity of the guanine quenching method using a masking DNA oligonucleotide. The simple addition of a masking DNA oligonucleotide was found to mask the interference of a different guanine in a target oligonucleotide on fluorescence and to enhance difference in the quenching ratio between wild-type and mutant oligonucleotides. Based on this strategy, we succeeded in discriminating various mutations from the wild-type YMDD motif of the hepatitis B virus DNA polymerase gene using guanine quenching with a masking oligonucleotide. 相似文献
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Shin KH Bae SD Hong HS Kim RH Kang MK Park NH 《Biochemical and biophysical research communications》2011,404(4):932-902
MicroRNAs (miRNAs) are epigenetic regulators of gene expression, and their deregulation plays an important role in human cancer, including oral squamous cell carcinoma (OSCC). Recently, we found that miRNA-181a (miR-181a) was upregulated during replicative senescence of normal human oral keratinocytes. Since senescence is considered as a tumor suppressive mechanism, we thus investigated the expression and biological role of miR-181a in OSCC. We found that miR-181a was frequently downregulated in OSCC. Ectopic expression of miR-181a suppressed proliferation and anchorage independent growth ability of OSCC. Moreover, miR-181a dramatically reduces the growth of OSCC on three dimensional organotypic raft culture. We also identified K-ras as a novel target of miR-181a. miR-181a decreased K-ras protein level as well as the luciferase activity of reporter vectors containing the 3′-untranslated region of K-ras gene. Finally, we defined a minimal regulatory region of miR-181a and found a positive correlation between its promoter activity and the level of miR-181a expression. In conclusion, miR-181a may function as an OSCC suppressor by targeting on K-ras oncogene. Thus, miR-181a should be considered for therapeutic application for OSCC. 相似文献
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Schulte G Bryja V Rawal N Castelo-Branco G Sousa KM Arenas E 《Journal of neurochemistry》2005,92(6):1550-1553
The Wnt family of lipoproteins regulates several aspects of the development of the nervous system. Recently, we reported that Wnt-3a enhances the proliferation of midbrain dopaminergic precursors and that Wnt-5a promotes their differentiation into dopaminergic neurones. Here we report the purification of hemagglutinin-tagged Wnt-5a using a three-step purification method similar to that previously described for Wnt-3a. Haemagglutinin-tagged Wnt-5a was biologically active and induced the differentiation of immature primary midbrain precursors into tyrosine hydroxylase-positive dopaminergic neurones. Using a substantia nigra-derived dopaminergic cell line (SN4741), we found that Wnt-5a, unlike Wnt-3a, did not promote beta-catenin phosphorylation or stabilization. However, both Wnt-5a and Wnt-3a activated dishevelled, as assessed by a phosphorylation-dependent mobility shift. Moreover, the activity of Wnt-5a on dishevelled was blocked by pre-treatment with acyl protein thioesterase-1, indicating that palmitoylation of Wnt-5a is necessary for its function. Thus, our results suggest that Wnt-3a and Wnt-5a, respectively, activate canonical and non-canonical Wnt signalling pathways in ventral midbrain dopaminergic cells. Furthermore, we identify dishevelled as a key player in transducing both Wnt canonical and non-canonical signals in dopaminergic cells. 相似文献
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Koduru S Massaad M Wilbur C Kumar L Geha R Ramesh N 《Biochemical and biophysical research communications》2007,353(4):875-881
The majority of Wiskott-Aldrich syndrome protein (WASP) in T cells is in a complex with WASP interacting protein (WIP), a 503 a.a. long proline rich protein. Here we demonstrate that a novel anti-WIP mAb, 3D10, recognizes an epitope in the N-terminal domain of the WIP protein, within the sequence 13PTFALA18. mAb 3D10 competes with actin, but not with WASP or Nck, for WIP binding. Analysis of 3D10 immunoprecipitates failed to demonstrate dissociation of the WASP-WIP complex after TCR ligation that we previously reported using a polyclonal anti-WIP anti-serum raised against a C-terminal peptide (a.a. 459-503) that spanned the WASP binding site. 3D10 mAb allowed the detection of a novel isoform of WIP consisting of a truncated 403 a.a. long protein that includes the 377 a.a. encoded by the first 4 exons of WIP followed by a 26 a.a. sequence encoded by intron 4. 相似文献