Human-Soybean Allergies: Elucidation of the Seed Proteome and Comprehensive Protein-Protein Interaction Prediction.

The soybean crop, Glycine max (L.) Merr., is consumed by humans, Homo sapiens, worldwide. While the respective bodies of literature and -omics data for each of these organisms are extensive, comparatively few studies investigate the molecular biological processes occurring between the two. We are interested in elucidating the network of protein-protein interactions (PPIs) involved in human-soybean allergies. To this end, we leverage state-of-the-art sequence-based PPI predictors amenable to predicting the enormous comprehensive interactome between human and soybean. A network-based analytical approach is proposed, leveraging similar interaction profiles to identify candidate allergens and proteins involved in the allergy response. Interestingly, the predicted interactome can be explored from two complementary perspectives: which soybean proteins are predicted to interact with specific human proteins and which human proteins are predicted to interact with specific soybean proteins. A total of eight proteins (six specific to the human proteome and two to the soy proteome) have been identified and supported by the literature to be involved in human health, specifically related to immunological and neurological pathways. This study, beyond generating the most comprehensive human-soybean interactome to date, elucidated a soybean seed interactome and identified several proteins putatively consequential to human health.

PIPE4: Fast PPI Predictor for Comprehensive Inter- and Cross-Species Interactomes.

The need for larger-scale and increasingly complex protein-protein interaction (PPI) prediction tasks demands that state-of-the-art predictors be highly efficient and adapted to inter- and cross-species predictions. Furthermore, the ability to generate comprehensive interactomes has enabled the appraisal of each PPI in the context of all predictions leading to further improvements in classification performance in the face of extreme class imbalance using the Reciprocal Perspective (RP) framework. We here describe the PIPE4 algorithm. Adaptation of the PIPE3/MP-PIPE sequence preprocessing step led to upwards of 50x speedup and the new Similarity Weighted Score appropriately normalizes for window frequency when applied to any inter- and cross-species prediction schemas. Comprehensive interactomes for three prediction schemas are generated: (1) cross-species predictions, where Arabidopsis thaliana is used as a proxy to predict the comprehensive Glycine max interactome, (2) inter-species predictions between Homo sapiens-HIV1, and (3) a combined schema involving both cross- and inter-species predictions, where both Arabidopsis thaliana and Caenorhabditis elegans are used as proxy species to predict the interactome between Glycine max (the soybean legume) and Heterodera glycines (the soybean cyst nematode). Comparing PIPE4 with the state-of-the-art resulted in improved performance, indicative that it should be the method of choice for complex PPI prediction schemas.

Genetic Analysis of High Protein Content in 'AC Proteus' Related Soybean Populations Using SSR, SNP, DArT and DArTseq Markers.

Key message: Several AC Proteus derived genomic regions (QTLs, SNPs) have been identified which may prove useful for further development of high yielding high protein cultivars and allele-specific marker developments. High seed protein content is a trait which is typically difficult to introgress into soybean without an accompanying reduction in seed yield. In a previous study, 'AC Proteus' was used as a high protein source and was found to produce populations that did not exhibit the typical association between high protein and low yield. Five high x low protein RIL populations and a high x high protein RIL population were evaluated by either quantitative trait locus (QTL) analysis or bulk segregant analyses (BSA) following phenotyping in the field. QTL analysis in one population using SSR, DArT and DArTseq markers found two QTLs for seed protein content on chromosomes 15 and 20. The BSA analyses suggested multiple genomic regions are involved with high protein content across the five populations, including the two previously mentioned QTLs. In an alternative approach to identify high protein genes, pedigree analysis identified SNPs for which the allele associated with high protein was retained in seven high protein descendants of AC Proteus on chromosomes 2, 17 and 18. Aside from the two identified QTLs (five genomic regions in total considering the two with highly elevated test statistic, but below the statistical threshold and the one with epistatic interactions) which were some distance from Meta-QTL regions and which were also supported by our BSA analysis within five populations. These high protein regions may prove useful for further development of high yielding high protein cultivars.

A global investigation of gene deletion strains that affect premature stop codon bypass in yeast, Saccharomyces cerevisiae.

Protein biosynthesis is an orderly process that requires a balance between rate and accuracy. To produce a functional product, the fidelity of this process has to be maintained from start to finish. In order to systematically identify genes that affect stop codon bypass, three expression plasmids, pUKC817, pUKC818 and pUKC819, were integrated into the yeast non-essential loss-of-function gene array (5000 strains). These plasmids contain three different premature stop codons (UAA, UGA and UAG, respectively) within the LacZ expression cassette. A fourth plasmid, pUKC815 that carries the native LacZ gene was used as a control. Transformed strains were subjected to large-scale ß-galactosidase lift assay analysis to evaluate production of ß-galactosidase for each gene deletion strain. In this way 84 potential candidate genes that affect stop codon bypass were identified. Three candidate genes, OLA1, BSC2, and YNL040W, were further investigated, and were found to be important for cytoplasmic protein biosynthesis.

Trimethylation enhancement using diazomethane (TrEnDi): rapid on-column quaternization of peptide amino groups via reaction with diazomethane significantly enhances sensitivity in mass spectrometry analyses via a fixed, permanent positive charge.

Defining cellular processes relies heavily on elucidating the temporal dynamics of proteins. To this end, mass spectrometry (MS) is an extremely valuable tool; different MS-based quantitative proteomics strategies have emerged to map protein dynamics over the course of stimuli. Herein, we disclose our novel MS-based quantitative proteomics strategy with unique analytical characteristics. By passing ethereal diazomethane over peptides on strong cation exchange resin within a microfluidic device, peptides react to contain fixed, permanent positive charges. Modified peptides display improved ionization characteristics and dissociate via tandem mass spectrometry (MS(2)) to form strong a2 fragment ion peaks. Process optimization and determination of reactive functional groups enabled a priori prediction of MS(2) fragmentation patterns for modified peptides. The strategy was tested on digested bovine serum albumin (BSA) and successfully quantified a peptide that was not observable prior to modification. Our method ionizes peptides regardless of proton affinity, thus decreasing ion suppression and permitting predictable multiple reaction monitoring (MRM)-based quantitation with improved sensitivity.

Recent advances in protein-protein interaction prediction: experimental and computational methods.

INTRODUCTION: Proteins within the cell act as part of complex networks, which allow pathways and processes to function. Therefore, understanding how proteins interact is a significant area of current research. AREAS COVERED: This review aims to present an overview of key experimental techniques (yeast two-hybrid, tandem affinity purification and protein microarrays) used to discover protein-protein interactions (PPIs), as well as to briefly discuss certain computational methods for predicting protein interactions based on gene localization, phylogenetic information, 3D structural modeling or primary protein sequence data. Due to the large-scale applicability of primary sequence-based methods, the authors have chosen to focus on this strategy for our review. There is an emphasis on a recent algorithm called Protein Interaction Prediction Engine (PIPE) that can predict global PPIs. The readers will discover recent advances both in the practical determination of protein interaction and the strategies that are available to attempt to anticipate interactions without the time and costs of experimental work. EXPERT OPINION: Global PPI maps can help understand the biology of complex diseases and facilitate the identification of novel drug target sites. This study describes different techniques used for PPI prediction that we believe will significantly impact the development of the field in a new future. We expect to see a growing number of similar techniques capable of large-scale PPI predictions.