Dataset Information

Characterization of a Complex Mixture of Immunomodulator Peptides Obtained from Autologous Urine.

ABSTRACT: A complex mixture of peptides plays a key role in the regulation of the immune system; different sources as raw materials mainly from animals and vegetables have been reported to provide these extracts. The batch-to-batch product consistency depends on in-process controls established. However, when an immunomodulator is a customized product obtained from the same volunteer who will receive the product to personalize the treatment, the criteria to establish the consistency between volunteers are different. In this sense, it is expected to have the same molecular weight range although the profile of peptide abundance is different. Here, we characterized the peptide profile of three extracts of an immunomodulator obtained from the urine of different volunteers suffering from three different diseases (i.e., allergic rhinitis, rheumatoid arthritis, and chronic rhinopharyngitis), using size exclusion chromatography (SEC) and mass spectrometry (MS). The peptides contained in the immunomodulators were stable after six months, stored in a refrigerator. Our results showed a chromatographic profile with the same range of low molecular weight (less than 17?kDa) in all analyzed samples by SEC; these results were also confirmed by MS showing an exact mass spectrum from 3 to 13?kDa. The fact that the peptide profiles were conserved during a six-month period at refrigeration conditions (2 to 8°C) maintaining the quality and stability of the immunomodulator supports the notion that it might be an alternative in the treatment of chronic hypersensibility disorders.

SUBMITTER: Fragoso A

PROVIDER: S-EPMC7154977 | biostudies-literature | 2020

REPOSITORIES: biostudies-literature

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Characterization of a Complex Mixture of Immunomodulator Peptides Obtained from Autologous Urine.

Fragoso Alberto A Pedraza-Jiménez Mérida M Espinoza-González Laura L Ceja-Mendoza María Luisa ML Sánchez-Mercado Hugo H Robles-Pérez Gloria G Granados Julio J Medina-Rivero Emilio E

Journal of immunology research 20200402

A complex mixture of peptides plays a key role in the regulation of the immune system; different sources as raw materials mainly from animals and vegetables have been reported to provide these extracts. The batch-to-batch product consistency depends on in-process controls established. However, when an immunomodulator is a customized product obtained from the same volunteer who will receive the product to personalize the treatment, the criteria to establish the consistency between volunteers are ...[more]

PMID: 32322594

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Project description:Natural and non-natural cyclic peptides are a crucial component in drug discovery programs because of their considerable pharmaceutical properties. Cyclosporin, microcystins, and nodularins are all notable pharmacologically important cyclic peptides. Because these biologically active peptides are often biosynthesized nonribosomally, they often contain nonstandard amino acids, thus increasing the complexity of the resulting tandem mass spectrometry data. In addition, because of the cyclic nature, the fragmentation patterns of many of these peptides showed much higher complexity when compared to related counterparts. Therefore, at the present time it is still difficult to annotate cyclic peptides MS/MS spectra. In this current work, an annotation program was developed for the annotation and characterization of tandem mass spectra obtained from cyclic peptides. This program, which we call MS-CPA is available as a web tool (http://lol.ucsd.edu/ms-cpa_v1/Input.py). Using this program, we have successfully annotated the sequence of representative cyclic peptides, such as seglitide, tyrothricin, desmethoxymajusculamide C, dudawalamide A, and cyclomarins, in a rapid manner and also were able to provide the first-pass structure evidence of a newly discovered natural product based on predicted sequence. This compound is not available in sufficient quantities for structural elucidation by other means such as NMR. In addition to the development of this cyclic annotation program, it was observed that some cyclic peptides fragmented in unexpected ways resulting in the scrambling of sequences. In summary, MS-CPA not only provides a platform for rapid confirmation and annotation of tandem mass spectrometry data obtained with cyclic peptides but also enables quantitative analysis of the ion intensities. This program facilitates cyclic peptide analysis, sequencing, and also acts as a useful tool to investigate the uncommon fragmentation phenomena of cyclic peptides and aids the characterization of newly discovered cyclic peptides encountered in drug discovery programs.

Project description:Background:The oat × maize addition (OMA) lines are used for mapping of the maize genome, the studies of centromere-specific histone (CENH3), gene expression, meiotic chromosome behavior and also for introducing maize C4 photosynthetic system to oat. The aim of our study was the identification and molecular-cytogenetic characterization of oat × maize hybrids. Methods:Oat DH lines and oat × maize hybrids were obtained using the wide crossing of Avena sativa L. with Zea mays L. The plants identified as having a Grande-1 retrotransposon fragment, which produced seeds, were used for genomic in situ hybridization (GISH). Results:A total of 138 oat lines obtained by crossing of 2,314 oat plants from 80 genotypes with maize cv. Waza were tested for the presence of maize chromosomes. The presence of maize chromatin was indicated in 66 lines by amplification of the PCR product (500 bp) generated using primers specific for the maize retrotransposon Grande-1. Genomic in situ hybridization (GISH) detected whole maize chromosomes in eight lines (40%). All of the analyzed plants possessed full complement of oat chromosomes. The number of maize chromosomes differed between the OMA lines. Four OMA lines possessed two maize chromosomes similar in size, three OMA-one maize chromosome, and one OMA-four maize chromosomes. In most of the lines, the detected chromosomes were labeled uniformly. The presence of six 45S rDNA loci was detected in oat chromosomes, but none of the added maize chromosomes in any of the lines carried 45S rDNA locus. Twenty of the analyzed lines did not possess whole maize chromosomes, but the introgression of maize chromatin in the oat chromosomes. Five of 66 hybrids were shorter in height, grassy type without panicles. Twenty-seven OMA lines were fertile and produced seeds ranging in number from 1-102 (in total 613). Sixty-three fertile DH lines, out of 72 which did not have an addition of maize chromosomes or chromatin, produced seeds in the range of 1-343 (in total 3,758). Obtained DH and OMA lines were fertile and produced seeds. Discussion:In wide hybridization of oat with maize, the complete or incomplete chromosomes elimination of maize occur. Hybrids of oat and maize had a complete set of oat chromosomes without maize chromosomes, and a complete set of oat chromosomes with one to four retained maize chromosomes.

Project description:There is a consistent demand for new biosensors for the detection of protein targets, and a systematic method for the rapid development of new sensors is needed. Here we present a platform where short unstructured peptides that bind to a desired target are selected using M13 phage display. The selected peptides are then chemically synthesized and immobilized on gold, allowing for detection of the target using electrochemical techniques such as electrochemical impedance spectroscopy (EIS). A quartz crystal microbalance (QCM) is also used as a diagnostic tool during biosensor development. We demonstrate the utility of this approach by creating a novel peptide-based electrochemical biosensor for the enzyme alanine aminotransferase (ALT), a well-known biomarker of hepatotoxicity. Biopanning of the M13 phage display library over immobilized ALT, led to the rapid identification of a new peptide (ALT5-8) with an amino acid sequence of WHWRNPDFWYLK. Phage particles expressing this peptide exhibited nanomolar affinity for immobilized ALT (K(d,app) = 85±20 nM). The newly identified ALT5-8 peptide was then chemically synthesized with a C-terminal cysteine for gold immobilization. The performance of the gold-immobilized peptides was studied with cyclic voltammetry (CV), QCM, and EIS. Using QCM, the sensitivity for ALT detection was 8.9±0.9 Hz/(µg/mL) and the limit of detection (LOD) was 60 ng/mL. Using EIS measurements, the sensitivity was 142±12 impedance percentage change %/(µg/mL) and the LOD was 92 ng/mL. In both cases, the LOD was below the typical concentration of ALT in human blood. Although both QCM and EIS produced similar LODs, EIS is preferable due to a larger linear dynamic range. Using QCM, the immobilized peptide exhibited a nanomolar dissociation constant for ALT (K(d) = 20.1±0.6 nM). These results demonstrate a simple and rapid platform for developing and assessing the performance of sensitive, peptide-based biosensors for new protein targets.

Dataset Information

Characterization of a Complex Mixture of Immunomodulator Peptides Obtained from Autologous Urine.

Publications

Characterization of a Complex Mixture of Immunomodulator Peptides Obtained from Autologous Urine.

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