scholarly journals Purified Box C/D snoRNPs Are Able To Reproduce Site-Specific 2′-O-Methylation of Target RNA In Vitro

2002 ◽  
Vol 22 (19) ◽  
pp. 6663-6668 ◽  
Author(s):  
Silvia Galardi ◽  
Alessandro Fatica ◽  
Angela Bachi ◽  
Andrea Scaloni ◽  
Carlo Presutti ◽  
...  
Keyword(s):  
Methylation Pattern ◽  
Spectrometric Analysis ◽  
Specific Factor ◽  
Site Specific ◽  
Protein Components ◽  
Target Rna ◽  
Nucleolar Rnas ◽  
Protein Tagging ◽  
Selection Of

ABSTRACT Small nucleolar RNAs (snoRNAs) are associated in ribonucleoprotein particles localized to the nucleolus (snoRNPs). Most of the members of the box C/D family function in directing site-specific 2′-O-methylation of substrate RNAs. Although the selection of the target nucleotide requires the antisense element and the conserved box D or D′ of the snoRNA, the methyltransferase activity is supposed to reside in one of the protein components. Through protein tagging of a snoRNP-specific factor, we purified to homogeneity box C/D snoRNPs from the yeast Saccharomyces cerevisiae. Mass spectrometric analysis demonstrated the presence of Nop1p, Nop58p, Nop56p, and Snu13p as integral components of the particle. We show that purified snoRNPs are able to reproduce the site-specific methylation pattern on target RNA and that the predicted S-adenosyl-l-methionine-binding region of Nop1p is responsible for the catalytic activity.

scholarly journals The central body of the cyanelles of Cyanophora paradoxa: a eukaryotic carboxysome?

2005 ◽  
Vol 83 (7) ◽  
pp. 758-764 ◽  
Author(s):  
S C Burey ◽  
S Fathi-Nejad ◽  
V Poroyko ◽  
J M Steiner ◽  
W Löffelhardt ◽  
...  
Keyword(s):  
Inorganic Carbon ◽  
Expressed Sequence Tag ◽  
Rubisco Activase ◽  
Spectrometric Analysis ◽  
Cyanophora Paradoxa ◽  
Carbon Concentrating Mechanism ◽  
High Accumulation ◽  
Concentrating Mechanism ◽  
Protein Components

The cyanelles of the glaucocystophyte Cyanophora paradoxa combine two prokaryotic features not found in other phototrophic eukaryotes: a peptidoglycan wall and a putative carboxysome. Both of them would be indispensable when a inorganic carbon concentrating mechanism involving high accumulation of bicarbonate in the cyanelle stroma is assumed. Two approaches were used. (i) An expressed sequence tag library was generated allowing access to interesting genes and microarray technology. Hybridization of the microarrays to RNA from cells grown at high and low CO2 yielded 97 genes that were upregulated under CO2 stress whereas 87 genes were found to be downregulated. (ii) Cyanelle central bodies were isolated and protein components other than Rubisco were investigated by mass spectrometry. So far, mass spectrometric analysis of putative carboxysomal proteins yielded only sequences with no match in the databases. Rubisco activase could be shown via in vitro import and Western blotting to be copackaged with Rubisco in isolated purified central bodies. While our data support the presence of an inorganic carbon concentrating mechanism in cyanelles, they do not allow us to distinguish the microcompartment as a carboxysome or pyrenoid.Key words: Cyanophora paradoxa, cyanelles, carboxysome, Rubisco activase, carbon-concentrating mechanism, microarrays.

Clone selection of in vitro horseradish hairy root cultures

Planta Medica ◽  
2015 ◽  
Vol 81 (16) ◽  
Author(s):  
R Bertóti ◽  
Á Alberti ◽  
A Böszörményi ◽  
R Könye ◽  
T Horváth ◽  
...  
Keyword(s):  
Hairy Root ◽  
Hairy Root Cultures ◽  
Root Cultures ◽  
Clone Selection ◽  
Selection Of

A Hexagonal (II) Phase Phospholipid Neutralization Assay for Lupus Anticoagulant Identification

1993 ◽  
Vol 70 (05) ◽  
pp. 787-793 ◽  
Author(s):  
Douglas A Triplett ◽  
Linda K Barna ◽  
Gail A Unger
Keyword(s):  
Hemophilia A ◽  
Clinical Laboratory ◽  
Neutralization Assay ◽  
Confirmatory Test ◽  
Specific Factor ◽  
Clinical Settings ◽  
Drug Induced ◽  
Variable Screening ◽  
Routine Clinical Laboratory

SummaryLupus anticoagulants (LAs) are immunoglobulins (IgG, IgM, or both) which interfere with in vitro phospholipid (PL) dependent tests of coagulation (e.g. APTT, dilute PT, dilute Russell Viper Venom Time). These antibodies may be identified in a wide variety of clinical settings. With the exception of heparinized patient samples, the presence of LAs is often the most common cause of an unexplained APTT in a routine clinical laboratory. The diagnosis of LAs is difficult due to variable screening reagent sensitivity and intrinsic heterogeneity of LAs. Recently, Rauch and colleagues have shown human monoclonal hybridoma LAs were inhibited by hexagonal (II) phase PLs. In contrast, lamellar phase PLs had no effect. We have evaluated a new assay system, Staclot LA®, which utilizes a hexagonal (II) phase PL (egg phosphatidylethanolamine [EPE]) as a confirmatory test for LAs. Plasma samples from the following patient populations were studied: LA positive, heparinized, oral anticoagulated, hemophilia A and B, and specific factor inhibitors (factors V, VIII, IX). Unlike previous studies, the LA positive patients were a mixed population including: autoimmune diseases, drug-induced, and post-infection. Our findings confirm the specificity of hexagonal (II) phase PL neutralization of LAs.

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