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81.
Linhui Zhai Cheng Chang Ning Li Duc M. Duong Hao Chen Zixin Deng Jian Yang Xuechuan Hong Yunping Zhu Ping Xu 《Proteomics》2013,13(15):2229-2237
Reversed phase microcolumns have been widely used for peptide pretreatment to desalt and remove interferences before tandem LC–MS in proteomics studies. However, few studies have characterized the effects of experimental parameters as well as column characteristics on the composition of identified peptides. In this study, several parameters including the concentration of ACN in washing buffer, the microcolumn's purification effect, the peptide recovery rate, and the dynamic‐binding capacity were characterized in detail, based upon stable isotope labeling by amino acids in a cell culture quantitative approach. The results showed that peptide losses can be reduced with low ACN concentration in washing buffers resulting in a recovery rate of approximately 82%. Furthermore, the effects of ACN concentration and loading amount on the properties of identified peptides were also evaluated. We found that the dynamic‐binding capacity of the column was approximately 26 μg. With increased loading amounts, more hydrophilic peptides were replaced by hydrophobic peptides. 相似文献
82.
Recently, we reported a novel proteomics quantitation scheme termed “combined precursor isotopic labeling and isobaric tagging (cPILOT)” that allows for the identification and quantitation of nitrated peptides in as many as 12–16 samples in a single experiment. cPILOT offers enhanced multiplexing and posttranslational modification specificity, however excludes global quantitation for all peptides present in a mixture and underestimates reporter ion ratios similar to other isobaric tagging methods due to precursor co‐isolation. Here, we present a novel chemical workflow for cPILOT that can be used for global tagging of all peptides in a mixture. Specifically, through low pH precursor dimethylation of tryptic or LysC peptides followed by high pH tandem mass tags, the same reporter ion can be used twice in a single experiment. Also, to improve triple‐stage mass spectrometry (MS3) data acquisition, a selective MS3 method that focuses on product selection of the y1 fragment of lysine‐terminated peptides is incorporated into the workflow. This novel cPILOT workflow has potential for global peptide quantitation that could lead to enhanced sample multiplexing and increase the number of quantifiable spectra obtained from MS3 acquisition methods. 相似文献
83.
双孢蘑菇子实体发育后期差异表达蛋白质分析 总被引:3,自引:2,他引:1
为探讨双孢蘑菇子实体发育后期的蛋白质表达变化,对双孢蘑菇As2796子实体采收期、成熟期和开伞期的蛋白质组进行了双向电泳(2-DE)分析,发现了16个表达差异明显的蛋白质。通过质谱分析(MALDI-TOF/TOF MS)和数据库检索,有14个差异蛋白质获得鉴定。其中磷酸烯醇式丙酮酸水合酶与能量代谢相关,T-蛋白复合体1、蛋白酶体、5-甲基四氢三谷氨酸-同型半胱氨酸甲基转移酶、1-吡咯琳-5-羧酸脱氢酶、精氨酸酶与氨基酸或蛋白质代谢直接相关,而GTP结合蛋白则参与细胞的多种生命活动,在细胞的生长发育过程中起着重要的作用。另外7个为功能未知的蛋白质。 相似文献
84.
Manousos Makridakis Maria G. Roubelakis Antonia Vlahou 《Biochimica et Biophysica Acta - Proteins and Proteomics》2013,1834(11):2380-2384
Stem cells have been considered as possible therapeutic vehicles for different health related problems such as cardiovascular and neurodegenerative diseases and cancer. Secreted molecules are key mediators in cell–cell interactions and influence the cross talk with the surrounding tissues. There is strong evidence supporting that crucial cellular functions such as proliferation, differentiation, communication and migration are strictly regulated from the cell secretome. The investigation of stem cell secretome is accumulating continuously increasing interest given the potential use of these cells in regenerative medicine. The scope of the review is to report the main findings from the investigation of stem cell secretome by the use of contemporary proteomics methods and discuss the current status of research in the field. This article is part of a Special Issue entitled: An Updated Secretome. 相似文献
85.
Sheila S. Jaswal 《Biochimica et Biophysica Acta - Proteins and Proteomics》2013,1834(6):1188-1201
Over the past two decades, hydrogen exchange mass spectrometry (HXMS) has achieved the status of a widespread and routine approach in the structural biology toolbox. The ability of hydrogen exchange to detect a range of protein dynamics coupled with the accessibility of mass spectrometry to mixtures and large complexes at low concentrations result in an unmatched tool for investigating proteins challenging to many other structural techniques. Recent advances in methodology and data analysis are helping HXMS deliver on its potential to uncover the connection between conformation, dynamics and the biological function of proteins and complexes. This review provides a brief overview of the HXMS method and focuses on four recent reports to highlight applications that monitor structure and dynamics of proteins and complexes, track protein folding, and map the thermodynamics and kinetics of protein unfolding at equilibrium. These case studies illustrate typical data, analysis and results for each application and demonstrate a range of biological systems for which the interpretation of HXMS in terms of structure and conformational parameters provides unique insights into function. This article is part of a Special Issue entitled: Mass spectrometry in structural biology. 相似文献
86.
87.
Many attempts on optimization of sorghum [Sorghum bicolor (L.)
Moench] tissue culture induction media have been made, but the culture system
remains with some bottlenecks compared to that of other crops. This study aimed
at assessing the suitability of various induction media to produce embryogenic
callus (yellow and friable) with high induction rates and reduced phenolic
exudation. The six culture medium modifications: 3 based on Murashige and
Skoog (MS) medium and one each based on Chu N6, Gamborg B5 and 190-2
media respectively were applied in the culture of mature embryos from 10
sorghum genotypes. Although there was a genotype influence on the attainment
of a yellow callus, friability of the callus was determined to be dependent on the
culture medium and not the genotype. Half strength MS medium with 0.2 mg/l
2,4-D with 2.8 g/l Gelrite® as the gelling agent modified with 1.0 g/l KH2PO4,
1.0 g/l L-proline, 1.0 g/l L-asparagine and 0.16 mg/l CuSO4·5H2O (type E) was
found to be the most effective resulting in about 60% yellow coloured callus
induction with 25% friability. Addition of CuSO4·5H2O, KH2PO4, L-proline and
L-asparagine significantly reduced the phenolic production. Half strength MS
medium was observed to contribute to quality callus production when compared
to full strength MS media modified with the compounds. The half strength MS
medium was also observed to suppress phenolic production. Medium 190-2
produced the highest regeneration frequency (40%) among the 3-regeneration
media tested. The results provide information on a suitable sorghum callus
induction medium necessary for embryogenesis. 相似文献
88.
The essential oil of the fresh leaves of Platycladus orientalis (L.), grown in four different biogeographic zones of Jordan,- (the Mediterranean, Irano-Turanian, Saharo-Arabian, and Sudanian penetration) -, were obtained by hydrodistillation and analysed by gas chromatography (GC) and gas chromatography-mass-spectrometry (GC/MS). The actual composition of the spontaneous emitted volatiles was obtained using the solid-phase-micro-extraction (SPME) method and investigated using the same chromatographic and spectroscopic methods. Hydrocarbon monoterpenes dominated the hydrodistilled oils and emissions of all regions. Bicyclic monoterpenes (sabinene, α-pinene, and α-thujene) and monocyclic α-terpinene were detected as the major constituents of the oils and emissions. Additionally, hierarchical cluster analysis (HCA) revealed that the clustering is based on the region of collection rather than the applied methodology. Differences were observed in the quantity of the obtained oils (P-values <0.01); the highest amount of volatile oil was obtained from samples grown in the Irano-Turanian biogeographic zone. 相似文献
89.
Harihar Milaganur Mohan Boning Yang Nicole A. Dean Malini Raghavan 《The Journal of biological chemistry》2020,295(49):16754
α1-antitrypsin (AAT) regulates the activity of multiple proteases in the lungs and liver. A mutant of AAT (E342K) called ATZ forms polymers that are present at only low levels in the serum and induce intracellular protein inclusions, causing lung emphysema and liver cirrhosis. An understanding of factors that can reduce the intracellular accumulation of ATZ is of great interest. We now show that calreticulin (CRT), an endoplasmic reticulum (ER) glycoprotein chaperone, promotes the secretory trafficking of ATZ, enhancing the media:cell ratio. This effect is more pronounced for ATZ than with AAT and is only partially dependent on the glycan-binding site of CRT, which is generally relevant to substrate recruitment and folding by CRT. The CRT-related chaperone calnexin does not enhance ATZ secretory trafficking, despite the higher cellular abundance of calnexin-ATZ complexes. CRT deficiency alters the distributions of ATZ-ER chaperone complexes, increasing ATZ-BiP binding and inclusion body formation and reducing ATZ interactions with components required for ER-Golgi trafficking, coincident with reduced levels of the protein transport protein Sec31A in CRT-deficient cells. These findings indicate a novel role for CRT in promoting the secretory trafficking of a protein that forms polymers and large intracellular inclusions. Inefficient secretory trafficking of ATZ in the absence of CRT is coincident with enhanced accumulation of ER-derived ATZ inclusion bodies. Further understanding of the factors that control the secretory trafficking of ATZ and their regulation by CRT could lead to new therapies for lung and liver diseases linked to AAT deficiency. 相似文献
90.