Universität zu Köln

Weng, Samuel S. H., Demir, Fatih ORCID: 0000-0002-5744-0205, Ergin, Enes K., Dirnberger, Sabrina, Uzozie, Anuli, Tuscher, Domenic, Nierves, Lorenz, Tsui, Janice, Huesgen, Pitter F. and Lange, Philipp F. (2019). Sensitive Determination of Proteolytic Proteoforms in Limited Microscale Proteome Samples. Mol. Cell. Proteomics, 18 (11). S. 2335 - 2348. ROCKVILLE: AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC. ISSN 1535-9484

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Abstract

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 mu g 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.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Weng, Samuel S. H. UNSPECIFIED UNSPECIFIED UNSPECIFIED Demir, Fatih UNSPECIFIED orcid.org/0000-0002-5744-0205 UNSPECIFIED Ergin, Enes K. UNSPECIFIED UNSPECIFIED UNSPECIFIED Dirnberger, Sabrina UNSPECIFIED UNSPECIFIED UNSPECIFIED Uzozie, Anuli UNSPECIFIED UNSPECIFIED UNSPECIFIED Tuscher, Domenic UNSPECIFIED UNSPECIFIED UNSPECIFIED Nierves, Lorenz UNSPECIFIED UNSPECIFIED UNSPECIFIED Tsui, Janice UNSPECIFIED UNSPECIFIED UNSPECIFIED Huesgen, Pitter F. UNSPECIFIED UNSPECIFIED UNSPECIFIED Lange, Philipp F. UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-129507
DOI:	10.1074/mcp.TIR119.001560
Journal or Publication Title:	Mol. Cell. Proteomics
Volume:	18
Number:	11
Page Range:	S. 2335 - 2348
Date:	2019
Publisher:	AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC
Place of Publication:	ROCKVILLE
ISSN:	1535-9484
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language N-TERMINAL PROTEOMICS; TRANSLATION INITIATION LANDSCAPE; PROTEASE; REVEALS; IDENTIFICATION; DEGRADATION; MATURATION; ENRICHMENT; PROJECT; TAILS Multiple languages Biochemical Research Methods Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/12950