Sense and Nonsense of Elevated Column Temperature in Proteomic Bottom-up LC-MS Analyses

Kód výsledku v IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F00179906%3A_____%2F21%3A10428644" target="_blank" >RIV/00179906:_____/21:10428644 - isvavai.cz</a>

Nalezeny alternativní kódy

RIV/00216208:11160/21:10428644

Výsledek na webu

<a href="https://verso.is.cuni.cz/pub/verso.fpl?fname=obd_publikace_handle&handle=5tj.0cK2ij" target="_blank" >https://verso.is.cuni.cz/pub/verso.fpl?fname=obd_publikace_handle&handle=5tj.0cK2ij</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1021/acs.jproteome.0c00479" target="_blank" >10.1021/acs.jproteome.0c00479</a>

Jazyk výsledku

angličtina

Název v původním jazyce

Sense and Nonsense of Elevated Column Temperature in Proteomic Bottom-up LC-MS Analyses

Popis výsledku v původním jazyce

Elevated column temperature represents a simple means for improving chromatographic separation of peptides. Here, we demonstrated the advantages of the column temperature in peptide separation using state-of-the-art columns. More importantly, we also determined how temperature can impair proteomic bottom-up analyses. We found that an elevated temperature in combination with the acidic pH of the mobile phase induced in-column peptide hydrolysis with high specificity to Asp and accelerated five modification reactions of amino acids. The positive effects of temperature dominated in the 30 min long gradients since the column operated at 90 degrees C provided the largest number of identified peptides and proteins. However, the adverse effects of temperature on peptide integrity in longer liquid chromatography-mass spectrometry (LC-MS) analyses required its reduction to obtain optimum results. The largest number of peptides was identified using the column maintained at 75 degrees C in 60 min long gradients, at 60 degrees C in 120 min long gradients, and at 45 degrees C in 240 min long gradients. Our results indicate that no universal column temperature exists for bottom-up LC-MS analyses. Quite the contrary, the temperature setting must be selected rationally to exploit the full capabilities of the state-of-the-art mass spectrometers in proteomic LC-MS analyses, with the gradient time being a critical factor.

Název v anglickém jazyce

Sense and Nonsense of Elevated Column Temperature in Proteomic Bottom-up LC-MS Analyses

Popis výsledku anglicky

Elevated column temperature represents a simple means for improving chromatographic separation of peptides. Here, we demonstrated the advantages of the column temperature in peptide separation using state-of-the-art columns. More importantly, we also determined how temperature can impair proteomic bottom-up analyses. We found that an elevated temperature in combination with the acidic pH of the mobile phase induced in-column peptide hydrolysis with high specificity to Asp and accelerated five modification reactions of amino acids. The positive effects of temperature dominated in the 30 min long gradients since the column operated at 90 degrees C provided the largest number of identified peptides and proteins. However, the adverse effects of temperature on peptide integrity in longer liquid chromatography-mass spectrometry (LC-MS) analyses required its reduction to obtain optimum results. The largest number of peptides was identified using the column maintained at 75 degrees C in 60 min long gradients, at 60 degrees C in 120 min long gradients, and at 45 degrees C in 240 min long gradients. Our results indicate that no universal column temperature exists for bottom-up LC-MS analyses. Quite the contrary, the temperature setting must be selected rationally to exploit the full capabilities of the state-of-the-art mass spectrometers in proteomic LC-MS analyses, with the gradient time being a critical factor.

Druh

J_imp - Článek v periodiku v databázi Web of Science

CEP obor

—

OECD FORD obor

30104 - Pharmacology and pharmacy

Projekt

<a href="/cs/project/EF15_003%2F0000465" target="_blank" >EF15_003/0000465: Vytvoření expertního týmu pro pokročilý výzkum v separačních vědách</a><br>

Návaznosti

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Rok uplatnění

2021

Kód důvěrnosti údajů

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

Název periodika

Journal of Proteome Research

ISSN

1535-3893

e-ISSN

—

Svazek periodika

20

Číslo periodika v rámci svazku

1

Stát vydavatele periodika

US - Spojené státy americké

Počet stran výsledku

13

Strana od-do

420-432

Kód UT WoS článku

000605145400037

EID výsledku v databázi Scopus

2-s2.0-85096050695