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Comparison of enrichment performance between HUNTER and TAILS

ABSTRACT: Protein N-termini reveal fundamental regulatory mechanisms and their perturbation in disease. We present a robust, sensitive, scalable and automatable method called High-efficiency Undecanal-based N Termini EnRichment (HUNTER) for system-wide identification of thousands of N-termini from minute samples. Comparison of enrichment performance between the HUNTER protocol and well-established TAILS protocol was based on HeLa cell lysates.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (ncbitaxon:9606)

SUBMITTER: Dr. Philipp Lange Dr. Pitter F. Huesgen

PROVIDER: MSV000084171 | MassIVE | Tue Aug 06 14:19:00 BST 2019

SECONDARY ACCESSION(S): PXD014931

REPOSITORIES: MassIVE

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Publications

Sensitive Determination of Proteolytic Proteoforms in Limited Microscale Proteome Samples.

Weng Samuel S H SSH Demir Fatih F Ergin Enes K EK Dirnberger Sabrina S Uzozie Anuli A Tuscher Domenic D Nierves Lorenz L Tsui Janice J Huesgen Pitter F PF Lange Philipp F PF

Molecular & cellular proteomics : MCP 20190830 11

Protein N termini unambiguously identify truncated, alternatively translated or modified proteoforms with distinct functions and reveal perturbations in disease. Selective enrichment of N-terminal peptides is necessary to achieve proteome-wide coverage for unbiased identification of site-specific regulatory proteolytic processing and protease substrates. However, many proteolytic processes are strictly confined in time and space and therefore can only be analyzed in minute samples that provide i ...[more]

PMID: 31471496

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Sensitive determination of proteolytic proteoforms in limited microscale proteome samples-Development of High-efficiency Undecanal-based N Termini EnRichment (HUNTER)

Project description:Protein N-termini reveal fundamental regulatory mechanisms and their perturbation in disease. We present a robust, sensitive, scalable and automatable method called High-efficiency Undecanal-based N Termini EnRichment (HUNTER) for system-wide identification of thousands of N-termini from minute samples. Optimization and automation of the HUNTER protocol was based on HeLa cell lysates and commercially-available plasma.

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Analysis of sorted human peripheral blood mononuclear cells by HUNTER (High-efficiency Undecanal-based N Termini EnRichment)

Project description:Protein N-termini reveal fundamental regulatory mechanisms and their perturbation in disease. We present a robust, sensitive, scalable and automatable method called High-efficiency Undecanal-based N Termini EnRichment (HUNTER) for system-wide identification of thousands of N-termini from minute samples. Identification of distinct N-terminal profiles in human peripheral blood mononuclear sorted cells (B-cells; NK; NKT; monocytes) as one of the applications of HUNTER.

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Analysis of mitochondrial N termini by HUNTER (High-efficiency Undecanal-based N Termini EnRichment)

Project description:Protein N-termini reveal fundamental regulatory mechanisms and their perturbation in disease. We present a robust, sensitive, scalable and automatable method called High-efficiency Undecanal-based N Termini EnRichment (HUNTER) for system-wide identification of thousands of N-termini from minute samples. Identification of distinct N-terminal profiles in mitochondrial-enriched B-ALL and AML samples as one of the applications of HUNTER. Mitochondria are enriched from 2.5 million cultured B-ALL or primary patient AML cells using pressure cycling (PCT) assisted lysis and differential density centrifugation.

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Optimization of HUNTER N-termini profiling using Arabidopsis thaliana Col-8 wt leaf proteome extracts

Project description:Optimization hydrophobic tagging reagent (hexadecanal or undecanal), reaction time (30 min to 16h) and evaluation of the HUNTER protocol performance with different amounts (25 ug to 400 ug) of purified Arabidopsis thaliana Col-8 wt leaf proteome as starting material.

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Top-Down proteomics and Bottom-Up terminomics for the Detection of Protein Termini in C. elegans

Project description:Here we investigate how top-down proteomics (TDP) can be of value for the detection of protein termini. Either GELFrEE or solid-phase enrichment was conducted for pre-fraction. To verify the detected N-termini HUNTER (High-efficiency Undecanal-based N Termini EnRichment) was employed. Reductive dimethylation on intact protein level was additionally applied.

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Project description:Arabidopsis thaliana Legumain (aka VPE) beta / gamma were characterized for their cleavage specificity and putative substrates were identified by the N-termini enrichment method HUNTER.

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Project description:Comparison of protein N-termini in Arabidopsis thaliana leaves by the HUNTER method; the corresponding digestion was performed with legumain, GluC and trypsin.

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Complete Genomes of a Hunter-Gatherer and a Bantu from Southern Africa

Project description:The genetic structure of the indigenous hunter-gatherer peoples of Southern Africa, the oldest known lineage of modern man, holds an important key to understanding humanity's early history. Previously sequenced human genomes have been limited to recently diverged populations. Here we present the first complete genome sequences of an indigenous hunter-gatherer from the Kalahari Desert and of a Bantu from Southern Africa, as well as protein-coding regions from an additional three hunter-gatherers from disparate regions of the Kalahari. We characterize the extent of whole-genome and exome diversity among the five men, reporting 1.3 million novel DNA differences genome-wide, and 13,146 novel amino-acid variants. These data allow genetic relationships among Southern African foragers and neighboring agriculturalists to be traced more accurately than was previously possible. Adding the described variants to current databases will facilitate inclusion of Southern Africans in medical research efforts.

2010-02-18 | GSE19048 | GEO

Natural selection has contributed to functional immune response differences between human hunter-gatherers and agriculturalists

Project description:The shift from a hunter-gatherer (HG) to an agricultural (AG) mode of subsistence is believed to have been associated with profound changes in the burden and diversity of pathogens across human populations. Yet, the extent to which the advent of agriculture impacted the evolution of the human immune system remains unknown. Here we present a comparative study of variation in the transcriptional responses of peripheral blood mononuclear cells (PBMCs) to bacterial and viral stimuli between the Batwa, a rainforest hunter-gatherer, and the Bakiga, an agriculturalist population from Central Africa. We observed increased divergence between hunter-gatherers and farmers in the transcriptional response to viruses compared to that for bacterial stimuli. We demonstrate that a significant fraction of these transcriptional differences are under genetic control, and we show that positive natural selection has helped to shape population differences in immune regulation. Unexpectedly, we found stronger signatures of recent natural selection in the rainforest hunter-gatherers, which argues against the popularized notion that shifts in pathogen exposure due to the advent of agriculture imposed radically heightened selective pressures in agriculturalist populations.

2018-09-27 | GSE120502 | GEO

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