Journal of Integrated -OMICS
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Adaptive-BLAST: A User-defined Platform for the Study of Proteins: DOI: 10.5584/jiomics.v1i1.33
No full text Profile-based protein-sequence analysis algorithms comprise some of the most powerful and user-friendly methods for exploring protein sequences to determine their structure, function, and/or evolution (1-4). PSI-BLAST (5, 6) and rps-BLAST (7) are two of the most popular profile-based algorithms (~1120 references to date), and have exceptional utility in the identification of homology between proteins, particularly for biological scientists who do not specialize in computational approaches. However, when the performance of these algorithms is compared to other methods [e.g. support-vector machine learning (SVM) (8), hidden-markov models (HMMs) (9)], they often underperform in identifying the aforementioned protein properties (3, 9-11). We have previously demonstrated that the utility of BLAST algorithms can be significantly improved by: (i) adaptations to the profile libraries employed, (ii) adjustments to output formats, and (iii) alterations to BLAST algorithm itself (4, 6, 12-14). We present here Adaptive-BLAST (Ada-BLAST), which provides a simple user-defined platform for measuring and analyzing primary amino acid sequences. Within this platform, we developed a series of local BLAST applications (apps) that take advantage of the speed and sensitivity afforded by BLAST, while allowing for maximal user-definitions and flexible visualization. We tested the efficacy of these apps in control experiments, studying fold-recognition, in which we obtained >90% accuracy in highly divergent sequences (>25% identity). In addition, these same apps were proficient in classifying transmembrane proteins, identifying structural/functional determinants of ion-channels/receptors, and informing structural modeling algorithms. Indeed, these Ada-BLAST informed-structural models were useful in guiding our experimental research on the N-terminus of Transient Receptor Potential ion-channels (TRPs). Taken together, we propose that Ada-BLAST provides a powerful computational tool that is accessible to bench-scientists and computational biologists alike. 
Proteomic evaluation of Escherichia coli isolates from human clinical strains: DOI: 10.5584/jiomics.v1i1.20
No full textAcquired resistance to beta-lactams is mainly mediated by extended-spectrum beta-lactamases (ESBLs) that confer bacterial resistance to all beta-lactams except carbapenems and cephamycins, which are inhibited by other beta-lactamase inhibitors such as clavulanic acid. Although ESBLs still constitute the first cause of resistance to beta-lactams among Escherichia coli, other “new beta-lactamases” conferring resistance to carbapenems, such as metallo-beta-lactamases (MBL) and KPC carbapenemases, or to cephamycins, such as CMY enzymes, have more recently emerged and are often associated with ESBLs. In order to identify and characterize the proteome of extended-spectrum β-lactamase (ESBL) type TEM-52 and CMY-2 producing-Escherichia coli strains of human clinical origin a bidimensional electrophoresis (2-DE) technique with an isoelectric focusing followed by a SDS-PAGE, were used. Full proteomic studies were conducted in the same IEF and SDS-PAGE conditions, for two protein samples of E. colistrains with similar antibiotic-resistance profiles recovered from human clinical sources. A total of 64 and 91 spots were recovered and identified in C583 and C580 strains, respectively. Our results will be helpful for further understanding of antibiotic-resistant mechanism
Application of high content biology demonstrates differential responses of keratin acetylation sites to short chain fatty acids and to mitosis: DOI: 10.5584/jiomics.v1i2.57
No full textThe intermediate filament cytoskeleton in epithelial tissues is formed of keratin heterodimers. Keratins are highly post-translationally modified proteins, with tyrosine phopsphorylation, serine phosphorylation, and glycosylation amongst reported modifications. We and others have recently reported multiple acetylation sites on keratin 8 and we have previously shown that these sites are responsive to butyrate. In this study, we report the application of cellomic approaches to demonstrate differential responses of three lysine acetylations (lys 10, lys 471 and lys 482) to different short-chain fatty acids. The data imply no fixed hierarchy of acetylation on keratin 8, and furthermore imply different ranges of HDAC inhibitory specificities for SCFA. Furthermore we have used the functionality of the HCA platform to show that the acetylation sites are differentially modified in cells undergoing mitosis. Taken together the data imply distinct roles for keratin acetylations in function, perhaps suggesting a keratin code
A method for estimation of immunogenic determinants mutability: case studies of HIV1 gp120 and diphtheria toxin: DOI: 10.5584/jiomics.v1i2.64
No full textThere is a need for the method which helps to choose the less mutable immunogenic determinant for the design of recombinant or synthetic vaccines and ELISA test-systems. With the help of our method based on the directional mutational pressure theory one is able to estimate direction of symmetric and asymmetric mutational pressure in a gene coding for a protein of interest, and to find out which of its immunogenic determinants are less prone to missense mutations and so, to immune escaping. Three original computer algorithms (“VVK Sliding Window”, “VVK VarInvar” and “VVK Protective Buffer” available via www.barkovsky.hotmail.ru) have been created to perform all the necessary calculations and tests. “VVK Sliding Window” calculates nucleotide usage in fourfold and twofold degenerated sites, as well as usage of missense, nonsense and synonymous sites for each kind of nucleotide mutation along the length of a coding region, while “VVK Protective Buffer” calculates those indexes in a set of sequences. “VVK VarInvar” calculates percentage of variable sites in a set of aligned sequences, as well as nucleotide usage in invariable sites. We tested our method on HIV1 gp120 protein and on diphtheria toxin. The less mutable epitopes have been found for both proteins. Finally, we showed that antibodies recognizing the less mutable epitope of gp120 can be found in 80,22% of HIV1-infected persons
Assessing the Loss of Information through Application of the ‘Two-hit Rule’ in iTRAQ Datasets: DOI: 10.5584/jiomics.v1i1.53
No full textHigh-throughput studies of complex protein mixtures using proteomic workflows typically employ tandem mass spectrometric analysis of peptides obtained by tryptic digestion. Protein identification is achieved by comparing the experimentally obtained peptide MS/MS spectra to theoretical spectra. Protein identifications based on peptide fragment sequences are often judged valid using the so called ‘two-peptide’ rule whereby any protein identified by sequencing of fragment ions must be justified by the identification of two sequence unique peptides from the same protein. This excludes proteins identified on the basis of a single peptide ‘hit’ (often termed a one-hit wonder, or OHW). Applying the ‘two hit’ stringency may result in the loss of potentially valuable meta-data: information yielded or consolidated by valid OHW proteins may be overlooked. This study tests the hypothesis that certain groups of OHW proteins (and thus related biological events or pathways) are more likely to be identified by single peptide due to various physical or biochemical characteristics (molecular weight and isoelectric point). We have undertaken analysis on data from three independent quantitative iTRAQ based proteomic studies of a human colon cell line and human colon tissue to correlate the differences between OHW and “valid” protein sets for molecular weight, isoelectric point and for associated biological pathways. The results show that there is a possible trend of inverse correlation between the pI value of a protein and the number of peptide hits for identification. Molecular weights range from 30-60 kDa. Pathway analysis using EBI-EMBL Reactome SkyPainter found that by excluding OHWs, several biological pathways were consistently not mapped, suggesting that exclusion of OHW potentially limits the understanding the biological processes potentially identified within the whole dataset. Future work should address strategies for evaluation of validity and reproducibility of these conclusions in other tissues