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J Proteome Res. 2017 Jun 2;16(6):2213-2220. doi: 10.1021/acs.jproteome.7b00005. Epub 2017 May 3.

HILAQ: A Novel Strategy for Newly Synthesized Protein Quantification.

Author information

1
Department of Chemical Physiology and Molecular and Cellular Neurobiology, The Scripps Research Institute , La Jolla, California 92037, United States.
2
Cambridge Isotope Laboratories, Inc. , 50 Frontage Road, Andover, Massachusetts 01810, United States.

Abstract

Here we describe a new strategy, HILAQ (Heavy Isotope Labeled Azidohomoalanine Quantification), to rapidly quantify the molecular vulnerability profile to oxytosis, which is an oxidative stress-induced programed cell death pathway that has been reported to be involved in aging and neurodegenerative diseases. HILAQ was able to quantify 1962 newly synthesized proteins (NSPs) after 1 h of pulse labeling in HEK293T cell line, while 353 proteins were quantified using the previously published QuaNCAT protocol. HILAQ was successfully applied to the HT22 oxytosis model. 226 proteins were found to have a two-fold change in abundance, and 108 proteins were enriched in the cell death pathway, demonstrating the utility of HT22 cells as a tool to study the molecular details of cell death involved in neurodegenerative diseases. The HILAQ strategy simplifies the analysis of newly synthesized proteomes through the use of isobaric labels and achieves higher sensitivity than previously published methods.

KEYWORDS:

HT22; azidohomoalanine; heavy isotope; new synthesized proteins; oxytosis; proteomics

PMID:
28437088
PMCID:
PMC5718913
DOI:
10.1021/acs.jproteome.7b00005
[Indexed for MEDLINE]
Free PMC Article

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