A three-pronged 'Pitchfork' strategy enables an extensive description of the human membrane proteome and the identification of missing proteins

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388971%3A_____%2F19%3A00517774" target="_blank" >RIV/61388971:_____/19:00517774 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216208:11110/19:10397761 RIV/00216208:11310/19:10397761

Výsledek na webu

<a href="https://www.sciencedirect.com/science/article/pii/S1874391919301836?via%3Dihub" target="_blank" >https://www.sciencedirect.com/science/article/pii/S1874391919301836?via%3Dihub</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1016/j.jprot.2019.103411" target="_blank" >10.1016/j.jprot.2019.103411</a>

Jazyk výsledku

angličtina

Název v původním jazyce

A three-pronged 'Pitchfork' strategy enables an extensive description of the human membrane proteome and the identification of missing proteins

Popis výsledku v původním jazyce

Integral membrane proteins are under-represented in standard proteomic analyses, mostly because of their low expression and absence of trypsin-cleavage sites in their hydrophobic transmembrane segments. Novel and effective strategies for membrane proteomic analysis aim at soluble N-glycosylated segments of integral membrane proteins (CSC, SPEG, N-glyco-FASP) or selectively target the hydrophobic transmembrane alpha-helical segments employing chemical peptide cleavage by CNBr (hpTC). We combined a solid phase enrichment of glycopeptides (SPEG) with a transmembrane segment-oriented hpTC method and a standard 'detergent and trypsin' approach into a three-pronged 'Pitchfork' strategy to maximize the membrane proteome coverage in human lymphoma cells. This strategy enabled the identification of > 1200 integral membrane proteins from all cellular compartments, including 105 CD antigens, 24 G protein-coupled receptors, and 141 solute carrier transporters. The advantage of the combination lies in the complementarity of the methods. SPEG and hpTC target different sets of membrane proteins. HpTC provided identifications of proteins and peptides with significantly higher hydrophobicity compared to SPEG and detergent-trypsin approaches. Among all identified proteins, we observed 32 so-called 'missing proteins'. The Pitchfork strategy presented here is universally applicable and enables deep and fast description of membrane proteomes in only 3 LC-MS/MS runs per replicate. Significance: Integral membrane proteins (IMPs) are encoded by roughly a quarter of human coding genes. Their functions and their specific localization makes IMPs highly attractive drug targets. In fact, roughly half of the currently approved drugs in medicine target IMPs. Our knowledge of membrane proteomes is, however, limited. We present a new strategy for the membrane proteome analysis that combines three complementary methods targeting different features of IMPs.

Název v anglickém jazyce

A three-pronged 'Pitchfork' strategy enables an extensive description of the human membrane proteome and the identification of missing proteins

Popis výsledku anglicky

Integral membrane proteins are under-represented in standard proteomic analyses, mostly because of their low expression and absence of trypsin-cleavage sites in their hydrophobic transmembrane segments. Novel and effective strategies for membrane proteomic analysis aim at soluble N-glycosylated segments of integral membrane proteins (CSC, SPEG, N-glyco-FASP) or selectively target the hydrophobic transmembrane alpha-helical segments employing chemical peptide cleavage by CNBr (hpTC). We combined a solid phase enrichment of glycopeptides (SPEG) with a transmembrane segment-oriented hpTC method and a standard 'detergent and trypsin' approach into a three-pronged 'Pitchfork' strategy to maximize the membrane proteome coverage in human lymphoma cells. This strategy enabled the identification of > 1200 integral membrane proteins from all cellular compartments, including 105 CD antigens, 24 G protein-coupled receptors, and 141 solute carrier transporters. The advantage of the combination lies in the complementarity of the methods. SPEG and hpTC target different sets of membrane proteins. HpTC provided identifications of proteins and peptides with significantly higher hydrophobicity compared to SPEG and detergent-trypsin approaches. Among all identified proteins, we observed 32 so-called 'missing proteins'. The Pitchfork strategy presented here is universally applicable and enables deep and fast description of membrane proteomes in only 3 LC-MS/MS runs per replicate. Significance: Integral membrane proteins (IMPs) are encoded by roughly a quarter of human coding genes. Their functions and their specific localization makes IMPs highly attractive drug targets. In fact, roughly half of the currently approved drugs in medicine target IMPs. Our knowledge of membrane proteomes is, however, limited. We present a new strategy for the membrane proteome analysis that combines three complementary methods targeting different features of IMPs.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

10608 - Biochemistry and molecular biology

Projekt

Výsledek vznikl pri realizaci vícero projektů. Více informací v záložce Projekty.

Návaznosti

P - Projekt vyzkumu a vyvoje financovany z verejnych zdroju (s odkazem do CEP)

Rok uplatnění

2019

Kód důvěrnosti údajů

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Název periodika

Journal of Proteomics

ISSN

1874-3919

e-ISSN

—

Svazek periodika

204

Číslo periodika v rámci svazku

JUL 30

Stát vydavatele periodika

NL - Nizozemsko

Počet stran výsledku

10

Strana od-do

103411

Kód UT WoS článku

000475994800013

EID výsledku v databázi Scopus

2-s2.0-85067021763