Liquid Chromatography Electron Capture Dissociation Tandem Mass Spectrometry (LC-ECD-MS/MS) versus Liquid Chromatography Collision-induced Dissociation Tandem Mass Spectrometry (LC-CID-MS/MS) for the Identification of Proteins

Electron capture dissociation (ECD) offers many advantages over the more traditional fragmentation techniques for the analysis of peptides and proteins, although the question remains: How suitable is ECD for incorporation within proteomic strategies for the identification of proteins? Here, we compare LC-ECD-MS/MS and LC-CID-MS/MS as techniques for the identification of proteins.Experiments were performed on a hybrid linear ion trap–Fourier transform ion cyclotron resonance mass spectrometer. Replicate analyses of a six-protein (bovine serum albumin, apo-transferrin,lysozyme, cytochrome c, alcohol dehydrogenase, and β-galactosidase) tryptic digest were performed and the results analyzed on the basis of overall protein sequence coverage and sequence tag lengths within individual peptides. The results show that although protein coverage was lower for LC-ECDMS/MS than for LC-CID-MS/MS, LC-ECD-MS/MS resulted in longer peptide sequence tags,providing greater confidence in protein assignment

A common core RNP structure shared between the small nuclear box C/D RNPs and the spliceosomal U 4 snRNP.

AbstractThe box C/D snoRNAs function in directing 2′-O-methylation and/or as chaperones in the processing of ribosomal RNA. We show here that Snu13p (15.5kD in human), a component of the U4/U6.U5 tri-snRNP, is also associated with the box C/D snoRNAs. Indeed, genetic depletion of Snu13p in yeast leads to a major defect in RNA metabolism. The box C/D motif can be folded into a stem-internal loop-stem structure, almost identical to the 15.5kD binding site in the U4 snRNA. Consistent with this, the box C/D motif binds Snu13p/15.5kD in vitro. The similarities in structure and function observed between the U4 snRNP (chaperone for U6) and the box C/D snoRNPs raises the interesting possibility that these particles may have evolved from a common ancestral RNP

RNA secondary structure prediction from multi-aligned sequences

It has been well accepted that the RNA secondary structures of most functional non-coding RNAs (ncRNAs) are closely related to their functions and are conserved during evolution. Hence, prediction of conserved secondary structures from evolutionarily related sequences is one important task in RNA bioinformatics; the methods are useful not only to further functional analyses of ncRNAs but also to improve the accuracy of secondary structure predictions and to find novel functional RNAs from the genome. In this review, I focus on common secondary structure prediction from a given aligned RNA sequence, in which one secondary structure whose length is equal to that of the input alignment is predicted. I systematically review and classify existing tools and algorithms for the problem, by utilizing the information employed in the tools and by adopting a unified viewpoint based on maximum expected gain (MEG) estimators. I believe that this classification will allow a deeper understanding of each tool and provide users with useful information for selecting tools for common secondary structure predictions.Comment: A preprint of an invited review manuscript that will be published in a chapter of the book `Methods in Molecular Biology'. Note that this version of the manuscript may differ from the published versio

Recruiting Hard-to-Reach Subjects for Exercise Interventions: A Multi-Centre and Multi-Stage Approach Targeting General Practitioners and Their Community-Dwelling and Mobility-Limited Patients

The general practitioner (GP)’s practice appears to be an ideal venue for recruiting community-dwelling older adults with limited mobility. This study (Current Controlled Trials ISRCTN17727272) aimed at evaluating the recruiting process used for a multi-centre exercise intervention (HOMEfit). Each of six steps resulted in an absolute number of patients (N1–N6). Sex and age (for N4–N6) and reasons for dropping out were assessed. Patient database screening (N1–N3) at 15 GP practices yielded N1 = 5,990 patients aged 70 and above who had visited their GP within the past 6 months, N2 = 5,467 after exclusion of institutionalised patients, N3 = 1,545 patients eligible. Using a pre-defined limitation algorithm in order to conserve the practices’ resources resulted in N4 = 1,214 patients (80.3 ± 5.6 years, 68% female), who were then officially invited to the final assessment of eligibility at the GP’s practice. N5 = 434 patients (79.5 ± 5.4 years, 69% female) attended the practice screening (n = 13 of whom had not received an official invitation). Finally, N6 = 209 (79.8 ± 5.2 years, 74% female) were randomised after they were judged eligible and had given their written informed consent to participate in the randomised controlled trial (overall recruitment rate: 4.4%). The general strategy of utilising a GP’s practice to recruit the target group proved beneficial. The data and experiences presented here can help planners of future exercise-intervention studies

RNAalifold: improved consensus structure prediction for RNA alignments

<p>Abstract</p> <p>Background</p> <p>The prediction of a consensus structure for a set of related RNAs is an important first step for subsequent analyses. RNAalifold, which computes the minimum energy structure that is simultaneously formed by a set of aligned sequences, is one of the oldest and most widely used tools for this task. In recent years, several alternative approaches have been advocated, pointing to several shortcomings of the original RNAalifold approach.</p> <p>Results</p> <p>We show that the accuracy of RNAalifold predictions can be improved substantially by introducing a different, more rational handling of alignment gaps, and by replacing the rather simplistic model of covariance scoring with more sophisticated RIBOSUM-like scoring matrices. These improvements are achieved without compromising the computational efficiency of the algorithm. We show here that the new version of RNAalifold not only outperforms the old one, but also several other tools recently developed, on different datasets.</p> <p>Conclusion</p> <p>The new version of RNAalifold not only can replace the old one for almost any application but it is also competitive with other approaches including those based on SCFGs, maximum expected accuracy, or hierarchical nearest neighbor classifiers.</p

Physicochemical analysis of rotavirus segment 11 supports a 'modified panhandle' structure and not the predicted alternative tRNA-like structure (TRLS)

.Rotaviruses are a major cause of acute gastroenteritis, which is often fatal in infants. The viral genome consists of 11 double-stranded RNA segments, but little is known about their cis-acting sequences and structural elements. Covariation studies and phylogenetic analysis exploring the potential structure of RNA11 of rotaviruses suggested that, besides the previously predicted "modified panhandle" structure, the 5' and 3' termini of one of the isoforms of the bovine rotavirus UKtc strain may interact to form a tRNA-like structure (TRLS). Such TRLSs have been identified in RNAs of plant viruses, where they are important for enhancing replication and packaging. However, using tRNA mimicry assays (in vitro aminoacylation and 3'- adenylation), we found no biochemical evidence for tRNA-like functions of RNA11. Capping, synthetic 3' adenylation and manipulation of divalent cation concentrations did not change this finding. NMR studies on a 5'- and 3'-deletion construct of RNA11 containing the putative intra-strand complementary sequences supported a predominant panhandle structure and did not conform to a cloverleaf fold despite the strong evidence for a predicted structure in this conserved region of the viral RNA. Additional viral or cellular factors may be needed to stabilise it into a form with tRNA-like properties

PicXAA-R: Efficient structural alignment of multiple RNA sequences using a greedy approach

<p>Abstract</p> <p>Background</p> <p>Accurate and efficient structural alignment of non-coding RNAs (ncRNAs) has grasped more and more attentions as recent studies unveiled the significance of ncRNAs in living organisms. While the Sankoff style structural alignment algorithms cannot efficiently serve for multiple sequences, mostly progressive schemes are used to reduce the complexity. However, this idea tends to propagate the early stage errors throughout the entire process, thereby degrading the quality of the final alignment. For multiple protein sequence alignment, we have recently proposed PicXAA which constructs an accurate alignment in a non-progressive fashion.</p> <p>Results</p> <p>Here, we propose PicXAA-R as an extension to PicXAA for greedy structural alignment of ncRNAs. PicXAA-R efficiently grasps both folding information within each sequence and local similarities between sequences. It uses a set of probabilistic consistency transformations to improve the posterior base-pairing and base alignment probabilities using the information of all sequences in the alignment. Using a graph-based scheme, we greedily build up the structural alignment from sequence regions with high base-pairing and base alignment probabilities.</p> <p>Conclusions</p> <p>Several experiments on datasets with different characteristics confirm that PicXAA-R is one of the fastest algorithms for structural alignment of multiple RNAs and it consistently yields accurate alignment results, especially for datasets with locally similar sequences. PicXAA-R source code is freely available at: <url>http://www.ece.tamu.edu/~bjyoon/picxaa/</url>.</p

Target‐oriented habitat and wildlife management: estimating forage quantity and quality of semi‐natural grasslands with Sentinel‐1 and Sentinel‐2 data

Semi‐natural grasslands represent ecosystems with high biodiversity. Their conservation depends on the removal of biomass, for example, through grazing by livestock or wildlife. For this, spatially explicit information about grassland forage quantity and quality is a prerequisite for efficient management. The recent advancements of the Sentinel satellite mission offer new possibilities to support the conservation of semi‐natural grasslands. In this study, the combined use of radar (Sentinel‐1) and multispectral (Sentinel‐2) data to predict forage quantity and quality indicators of semi‐natural grassland in Germany was investigated. Field data for organic acid detergent fibre concentration (oADF), crude protein concentration (CP), compressed sward height (CSH) and standing biomass dry weight (DM) collected between 2015 and 2017 were related to remote sensing data using the random forest regression algorithm. In total, 102 optical‐ and radar‐based predictor variables were used to derive an optimized dataset, maximizing the predictive power of the respective model. High R2 values were obtained for the grassland quality indicators oADF (R2 = 0.79, RMSE = 2.29%) and CP (R2 = 0.72, RMSE = 1.70%) using 15 and 8 predictor variables respectively. Lower R2 values were achieved for the quantity indicators CSH (R2 = 0.60, RMSE = 2.77 cm) and DM (R2 = 0.45, RMSE = 90.84 g/m²). A permutation‐based variable importance measure indicated a strong contribution of simple ratio‐based optical indices to the model performance. In particular, the ratios between the narrow near‐infrared and red‐edge region were among the most important variables. The model performance for oADF, CP and CSH was only marginally increased by adding Sentinel‐1 data. For DM, no positive effect on the model performance was observed by combining Sentinel‐1 and Sentinel‐2 data. Thus, optical Sentinel‐2 data might be sufficient to accurately predict forage quality, and to some extent also quantity indicators of semi‐natural grassland

Fully three-dimensional sound speed-corrected multi-wavelength photoacoustic breast tomography

Photoacoustic tomography is a contrast agent-free imaging technique capable of visualizing blood vessels and tumor-associated vascularization in breast tissue. While sophisticated breast imaging systems have been recently developed, there is yet much to be gained in imaging depth, image quality and tissue characterization capability before clinical translation is possible. In response, we have developed a hybrid photoacoustic and ultrasound-transmission tomographic system PAM3. The photoacoustic component has for the first time three-dimensional multi-wavelength imaging capability, and implements substantial technical advancements in critical hardware and software sub-systems. The ultrasound component enables for the first time, a three-dimensional sound speed map of the breast to be incorporated in photoacoustic reconstruction to correct for inhomogeneities, enabling accurate target recovery. The results demonstrate the deepest photoacoustic breast imaging to date namely 48 mm, with a more uniform field of view than hitherto, and an isotropic spatial resolution that rivals that of Magnetic Resonance Imaging. The in vivo performance achieved, and the diagnostic value of interrogating angiogenesis-driven optical contrast as well as tumor mass sound speed contrast, gives confidence in the system's clinical potential.Comment: 33 pages Main Body, 9 pages Supplementary Materia