Dataset Information

Mass spectrometry of in-gel digests reveals differences in amino acid sequences of high-molecular-weight glutenin subunits in spelt and emmer compared to common wheat

ABSTRACT: High-molecular-weight glutenin subunits (HMW-GS) play an important role for the baking quality of wheat. The ancient wheats emmer and spelt differ in their HMW-GS pattern compared to modern common wheat and this might be one reason for their comparatively poor baking quality. The aim of this study was to elucidate similarities and differences in the amino acid sequences of two 1Bx HMW-GS of common wheat, spelt and emmer. First, the sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) system was optimized to separate common wheat, spelt and emmer Bx6 and Bx7 from other HMW-GS (e.g., 1Ax and 1By) in high concentrations. The in-gel digests of the Bx6 and Bx7 bands were analyzed by untargeted LC-MS/MS experiments revealing different UniProtKB accessions in spelt and emmer compared to common wheat. The HMW-GS Bx6 and Bx7, respectively, of emmer and spelt showed differences in the amino acid sequences compared to those of common wheat. The identities of the peptide variations were confirmed by targeted LC-MS/MS. These peptides can be used to differentiate between Bx6 and Bx7 of spelt and emmer and Bx6 and Bx7 of common wheat. The findings should help to increase the reliability and curation status of wheat protein databases and to understand the effects of protein structure on the functional properties.

ORGANISM(S): Triticum Dicoccoides Triticum Aestivum Triticum Spelta

SUBMITTER: Sabrina Geisslitz

PROVIDER: PXD016342 | panorama | Wed Jan 15 00:00:00 GMT 2020

REPOSITORIES: PanoramaPublic

ACCESS DATA

Publications

Mass spectrometry of in-gel digests reveals differences in amino acid sequences of high-molecular-weight glutenin subunits in spelt and emmer compared to common wheat.

Geisslitz Sabrina S America Antoine H P AHP Scherf Katharina Anne KA

Analytical and bioanalytical chemistry 20200111 6

High-molecular-weight glutenin subunits (HMW-GS) play an important role for the baking quality of wheat. The ancient wheats emmer and spelt differ in their HMW-GS pattern compared to modern common wheat and this might be one reason for their comparatively poor baking quality. The aim of this study was to elucidate similarities and differences in the amino acid sequences of two 1Bx HMW-GS of common wheat, spelt and emmer. First, the sodium dodecyl polyacrylamide gel electrophoresis (SDS-PAGE) sys ...[more]

PMID: 31927602

Similar Datasets

Project description:Glatiramer acetate (GA; Copaxone), is a complex mixture of synthetic polypeptides approved for treatment of multiple sclerosis (MS). GA is an altered peptide ligand (APL) of myelin basic protein (MBP), an encephalitic autoantigen implicated in MS. GA induces GA-reactive T cells, which upregulate expression of anti-inflammatory and neurotrophic substances in the CNS. The antigenic sequences in glatiramoids cannot be completely characterized; nevertheless, differences among them can cause toxicity. We conducted microarray analyses to determine whether gene expression by GA-reactive lymphocytes from mouse spleens could differentiate GA from other glatiramoids. Expression of 135 genes was unique to cells reactivated by GA vs by intentionally altered GA and other glatiramoids. Significantly (p<0.01) altered expression of 207 genes was observed by cells reactivated by GA vs purported generic GA. Some APLs of MBP have caused serious adverse effects in MS patients. The influence of altered gene expression on glatiramoid safety warrants further investigation. Glatiramoid samples for SPL cell activation were grouped into four categories: 1) Verified GA, which included GA-RS (22 samples) and GA drug product (GA-DP, 34 samples from 30 batches) manufactured by Teva; 2) Deliberately Modified GA (DM-GA; 9 samples), which included glatiramoids made by Teva that were similar to GA but modified in a variety of ways: prepared with different ingredients (e.g., missing a constituent amino acid); prepared with the same amino acids in the same molar ratio as GA, but with defined amino acid sequences and different molecular weights (referred to as peptide markers TV-35 and TV-66); synthesized by a different process (e.g., changing acetolytic cleavage conditions, alternating polymerization initiator); exposed to destabilizing conditions (e.g., degradation by acid, base, and heat); 3) Unverified Glatiramoids, which included 4 samples, TV-5010 and 3 glatiramoids synthesized to be similar to GA but not manufactured using the Teva-patented manufacturing process; and 4) Unverified Generic GA, which included samples from 2 glatiramoids (M-bM-^@M-^\GA-NM-bM-^@M-^] 11 samples from 5 different batches, and 2 M-bM-^@M-^\GA-CM-bM-^@M-^] samples from a single batch) marketed as generic GA manufactured by companies other than Teva .

Project description:Infiltrating T-lymphocytes from the peripheral blood into the central nervous system (CNS) play a dynamic role in the development of a neurological immune-mediated diseases. HAM/TSP is a chronic progressive inflammatory neurological disorder associated with human T-cell lymphotropic virus type I (HTLV-I) infection. In this chronic myelopathy, virus-infected circulating T-cells infiltrate the CNS and an immune response is initiated against the components of CNS. As the HTLV-I proviral load (PVL) has been used as the best clinical marker for patient diagnostic with HAM/TSP, we hypothesized there might be a signature on T-cell receptor (TCR) clonal repertoire in these patients, which could distinguish HAM/TSP patients from the healthy population, as well as from patients with a more heterogeneous CNS-reactive inflammatory disease as multiple sclerosis (MS). With this in mind, we applied an innovative unbiased molecular technique – unique molecular identifier (UMI) library-strategy to investigate with high accuracy the TCR clonal repertoire by high throughput sequencing (HTS) technology. cDNA-TCR β-chain libraries were sequenced from 2 million peripheral mononuclear cells (PBMCs) in 14 HAM/TSP patients, 34 MS patients and 20 healthy controls (HC). To address whether the clonal expansion correlates with the patient’s PVL level, analysis of longitudinal TCR repertoire was performed in 2 HAM/TSP patients. Over 5.6 million TCR sequences were generated per sample on HiSeq 2500 Illumina system and analyzed through the molecular identifier groups-based error correction pipeline (MiGEC). Bioinformatic analysis showed that clones with more than 8 reads had a lower coefficient of variation (CV) and then could be used with confidence to evaluate the TCR clonal expansion. While HAM/TSP patients showed the higher clonal T-cell expansion compared to MS and HC, increase of the TCR clonal expansion was inversely correlated with the diversity of TCR repertoire in all subject’s group. In addition, correlation of the PVL with TCR clonal expansion was observed in HAM/TSP patients at longitudinal time-points. Surprisingly, MS patients showed a higher diversity of TCR repertoire along with a very slight clonal T-cell expansion in comparison to either HAM/TSP patients or HC. Despite of the higher TCR clonal expansion in HAM/TSP patients, a non-shared or “private” TCR repertoire was observed in these patients. No clones that shared the same CDR3 amino acid sequences were seen in HC and MS patients. However, a cluster of related CDR3 amino acid sequences were observed for 18 out of 34 MS patients when evaluated by phylogenetic tree analysis. It suggestes that a TCR-repertoire signature might characterize patients with MS. Our findings suggest that even though a unique TCR-b repertoire shapes the immune response in patients with neurological immune-mediated disease, a relatedness on clonal T-cell repertoire exist in MS.

Dataset Information

Mass spectrometry of in-gel digests reveals differences in amino acid sequences of high-molecular-weight glutenin subunits in spelt and emmer compared to common wheat

Publications

Mass spectrometry of in-gel digests reveals differences in amino acid sequences of high-molecular-weight glutenin subunits in spelt and emmer compared to common wheat.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets