TY  - JOUR
AU  - Weng, Samuel SH
AU  - Demir, Fatih
AU  - Ergin, Enes K.
AU  - Dirnberger, Sabrina
AU  - Uzozie, Anuli
AU  - Tuscher, Domenic
AU  - Nierves, Lorenz
AU  - Tsui, Janice
AU  - Huesgen, Pitter F.
AU  - Lange, Philipp F.
TI  - Sensitive determination of proteolytic proteoforms in limited microscale proteome samples
JO  - Molecular & cellular proteomics
VL  - 18
IS  - 11
SN  - 1535-9484
CY  - Bethesda, Md.
PB  - The American Society for Biochemistry and Molecular Biology
M1  - FZJ-2019-04556
SP  - 2335-2347
PY  - 2019
AB  - 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 insufficient starting material for current enrichment protocols. Here we present High-efficiency Undecanal-based N Termini EnRichment (HUNTER), a robust, sensitive and scalable method for the analysis of previously inaccessible microscale samples. HUNTER achieved identification of >1000 N termini from as little as 2 μg raw HeLa cell lysate. Broad applicability is demonstrated by the first N-terminome analysis of sorted human primary immune cells and enriched mitochondrial fractions from pediatric cancer patients, as well as protease substrate identification from individual Arabidopsis thaliana wild type and Vacuolar Processing Enzyme-deficient mutant seedlings. We further implemented the workflow on a liquid handling system and demonstrate the feasibility of clinical degradomics by automated processing of liquid biopsies from pediatric cancer patients.
LB  - PUB:(DE-HGF)16
C6  - pmid:31471496
UR  - <Go to ISI:>//WOS:000494461000015
DO  - DOI:10.1074/mcp.TIR119.001560
UR  - https://juser.fz-juelich.de/record/864979
ER  -