Divergent-flow isoelectric focusing for separation and
preparative analysis of peptides

J 2012

Divergent-flow isoelectric focusing for separation and preparative analysis of peptides

DUŠA, Filip, Jana KŘENKOVÁ, Dana MORAVCOVÁ, Vladislav KAHLE, Karel ŠLAIS et. al.

Basic information

Original name

Divergent-flow isoelectric focusing for separation and preparative analysis of peptides

Authors

DUŠA, Filip (203 Czech Republic, belonging to the institution), Jana KŘENKOVÁ (203 Czech Republic), Dana MORAVCOVÁ (203 Czech Republic), Vladislav KAHLE (203 Czech Republic) and Karel ŠLAIS (203 Czech Republic)

Edition

Electrophoresis, Hoboken, WILEY-BLACKWELL, 2012, 0173-0835

Other information

Language

English

Type of outcome

Článek v odborném periodiku

Field of Study

10608 Biochemistry and molecular biology

Country of publisher

United States of America

Confidentiality degree

není předmětem státního či obchodního tajemství

References:

Full Text

Impact factor

Impact factor: 3.261

RIV identification code

RIV/00216224:14310/12:00081936

Organization unit

Faculty of Science

DOI

http://dx.doi.org/10.1002/elps.201100587

UT WoS

000305792500002

Keywords in English

Autofocusing; Caseinomacropeptide; Isoelectric focusing; Mass spectrometry; Peptides

Abstract

V originále

A divergent-flow isoelectric focusing (DF IEF) technique has been applied for the separation and preparative analysis of peptides. The parameters of the developed DF IEF device such as dimension and shape of the separation bed, selection of nonwoven material of the channel, and separation conditions were optimized. The DF IEF device was tested by the separation of a peptide mixture originating from the tryptic digestion of BSA, cytochrome c, and myoglobin. The pH gradient of DF IEF was created by the autofocusing of tryptic peptides themselves without any addition of carrier ampholytes. The focusing process was monitored visually using colored pI markers, and the obtained fractions were analyzed by RP-HPLC and ESI/TOF-MS. DF IEF operating in the autofocusing mode provides an efficient preseparation of peptides, which is comparable with a commercially available MicroRotofor multicompartment electrolyzer and significantly improves sequence coverage of analyzed proteins. The potential of the DF IEF device as an efficient tool for the preparative scale separations was demonstrated by the isolation of caseinomacropeptide (CMP) from a crude whey solution.

Links

IAAX00310701, research and development project

Name: Rychlá detekce a identifikace patogenních mikroorganizmů a virů pomocí elektromigračních technik a hmotnostní spektrometrie

Investor: Academy of Sciences of the Czech Republic

Citovat

DUŠA, Filip, Jana KŘENKOVÁ, Dana MORAVCOVÁ, Vladislav KAHLE and Karel ŠLAIS. Divergent-flow isoelectric focusing for separation and preparative analysis of peptides. Electrophoresis. Hoboken: WILEY-BLACKWELL, 2012, vol. 33, No 12, p. 1687-1694. ISSN 0173-0835. Available from: https://dx.doi.org/10.1002/elps.201100587.

@article{1138792,
   author = {Duša, Filip and Křenková, Jana and Moravcová, Dana and Kahle, Vladislav and Šlais, Karel},
   article_location = {Hoboken},
   article_number = {12},
   doi = {http://dx.doi.org/10.1002/elps.201100587},
   keywords = {Autofocusing; Caseinomacropeptide; Isoelectric focusing; Mass spectrometry; Peptides},
   language = {eng},
   issn = {0173-0835},
   journal = {Electrophoresis},
   title = {Divergent-flow isoelectric focusing for separation and preparative analysis of peptides},
   url = {https://onlinelibrary.wiley.com/doi/pdf/10.1002/elps.201100587},
   volume = {33},
   year = {2012}
}

TY  - JOUR
ID  - 1138792
AU  - Duša, Filip - Křenková, Jana - Moravcová, Dana - Kahle, Vladislav - Šlais, Karel
PY  - 2012
TI  - Divergent-flow isoelectric focusing for separation and preparative analysis of peptides
JF  - Electrophoresis
VL  - 33
IS  - 12
SP  - 1687-1694
EP  - 1687-1694
PB  - WILEY-BLACKWELL
SN  - 01730835
KW  - Autofocusing
KW  - Caseinomacropeptide
KW  - Isoelectric focusing
KW  - Mass spectrometry
KW  - Peptides
UR  - https://onlinelibrary.wiley.com/doi/pdf/10.1002/elps.201100587
L2  - https://onlinelibrary.wiley.com/doi/pdf/10.1002/elps.201100587
N2  - A divergent-flow isoelectric focusing (DF IEF) technique has been applied for the separation and preparative analysis of peptides. The parameters of the developed DF IEF device such as dimension and shape of the separation bed, selection of nonwoven material of the channel, and separation conditions were optimized. The DF IEF device was tested by the separation of a peptide mixture originating from the tryptic digestion of BSA, cytochrome c, and myoglobin. The pH gradient of DF IEF was created by the autofocusing of tryptic peptides themselves without any addition of carrier ampholytes. The focusing process was monitored visually using colored pI markers, and the obtained fractions were analyzed by RP-HPLC and ESI/TOF-MS. DF IEF operating in the autofocusing mode provides an efficient preseparation of peptides, which is comparable with a commercially available MicroRotofor multicompartment electrolyzer and significantly improves sequence coverage of analyzed proteins. The potential of the DF IEF device as an efficient tool for the preparative scale separations was demonstrated by the isolation of caseinomacropeptide (CMP) from a crude whey solution.
ER  -

DUŠA, Filip, Jana KŘENKOVÁ, Dana MORAVCOVÁ, Vladislav KAHLE and Karel ŠLAIS. Divergent-flow isoelectric focusing for separation and preparative analysis of peptides. \textit{Electrophoresis}. Hoboken: WILEY-BLACKWELL, 2012, vol.~33, No~12, p.~1687-1694. ISSN~0173-0835. Available from: https://dx.doi.org/10.1002/elps.201100587.

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