Dataset Information

Peptide recovery from acid-precipitated sodium deoxycholate pellet

ABSTRACT: Sodium deoxycholate (SDC) is MS-compatible ionic detergent which is compatible with in-solution tryptic digestion. SDC is removed from tryptic digested mixtures easily by acid precipitation and it has been demonstrated its capacity to increase trypsin activity. Additionally, and on the contrary to urea, SDC does not generate carbamylation side-products. Here, we report an optimized SDC-based shotgun proteomics sample preparation methods for blood-based protein biomarker research applied to human plasma. We show that an important subset of peptides was co-precipitated with the SDC salts and was recovered by a sequencial extraction of the SDC pellet by re-solubilization and precipitation.

INSTRUMENT(S): LTQ FT

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Blood Plasma

SUBMITTER: Aida Serra

LAB HEAD: Siu Kwan Sze

PROVIDER: PXD002811 | Pride | 2016-02-04

REPOSITORIES: Pride

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Publications

Plasma proteome coverage is increased by unique peptide recovery from sodium deoxycholate precipitate.

Serra Aida A Zhu Hongbin H Gallart-Palau Xavier X Park Jung Eun JE Ho Hee Haw HH Tam James P JP Sze Siu Kwan SK

Analytical and bioanalytical chemistry 20160125 7

The ionic detergent sodium deoxycholate (SDC) is compatible with in-solution tryptic digestion and LC-MS/MS-based shotgun proteomics by virtue of being easy to separate from the peptide products via precipitation in acidic buffers. However, it remains unclear whether unique human peptides co-precipitate with SDC during acid treatment of complex biological samples. In this study, we demonstrate for the first time that a large quantity of unique peptides in human blood plasma can be co-precipitate ...[more]

PMID: 26804737

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Project description:Here we exploit the essential process of X-chromosome dosage compensation to elucidate basic mechanisms that control the assembly, genome-wide binding, and function of gene regulatory complexes that act over large chromosomal territories. We demonstrate that a subunit of C. elegans MLL/COMPASS, a gene-activation complex, acts within the dosage compensation complex (DCC), a condensin complex, to target the DCC to both X chromosomes of hermaphrodites and thereby reduce chromosome-wide gene expression. The DCC binds to two categories of sites on X: rex sites that recruit the DCC in an autonomous, sequence- dependent manner, and dox sites that reside primarily in promoters of expressed genes and bind the DCC robustly only when attached to X. We find that DCC mutants that abolish rex-site binding do not eliminate dox-site binding, but instead reduce it to the level observed at autosomal binding sites in wild-type animals. Changes in DCC binding to these non-rex sites occur throughout development and correlate with transcriptional activity of adjacent genes. Moreover, autosomal DCC binding is enhanced by rex-site binding in cis in X-autosome fusion chromosomes. Thus, dox and autosomal sites exhibit similar binding properties. Our data support a model for DCC binding in which low-level DCC binding at dox and autosomal sites is dictated by intrinsic properties correlated with high transcriptional activity. Sex-specific DCC recruitment to rex sites then greatly elevates DCC binding to dox sites in cis, which lack intrinsically high DCC affinity on their own. We also show here that the C. elegans DCC achieves dosage compensation through its effects on transcription. ChIP-chip experiments using antibodies against DPY-27, SDC-3, DPY-30, DPY-26, MIX-1, SMC-4, ASH-2 in wild-type embryos. ChIP-chip experiments using antibodies against SDC-3, DPY-27, DPY-30, ASH-2, and IgG in different DCC mutants. ChIP-chip experiments using antibodies against RNA Pol II (hypophosphorylated and S2 and S5) in wild type and sdc-2 partial loss of function mutants.

Dataset Information

Peptide recovery from acid-precipitated sodium deoxycholate pellet

Publications

Plasma proteome coverage is increased by unique peptide recovery from sodium deoxycholate precipitate.

Similar Datasets

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