Enzyme kinetics above denaturation temperature: A temperature-jump/stopped- flow apparatus

Bálint Kintses, Zoltán Simon, Máté Gyimesi, Júlia Tóth, Balázs Jelinek, Csaba Niedetzky, M. Kovács, A. Málnási-Csizmadia

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

We constructed a "temperature-jump/stopped-flow" apparatus that allows us to study fast enzyme reactions at extremely high temperatures. This apparatus is a redesigned stopped-flow which is capable of mixing the reactants on a submillisecond timescale concomitant with a temperature-jump even as large as 60°C. We show that enzyme reactions that are faster than the denaturation process can be investigated above denaturation temperatures. In addition, the temperature-jump/ stopped-flow enables us to investigate at physiological temperature the mechanisms of many human enzymes, which was impossible until now because of their heat instability. Furthermore, this technique is extremely useful in studying the progress of heat-induced protein unfolding. The temperature-jump/stopped-flow method combined with the application of structure-specific fluorescence signals provides novel opportunities to study the stability of certain regions of enzymes and identify the unfoldinginitiating regions of proteins. The temperature-jump/stopped-flow technique may become a breakthrough in exploring new features of enzymes and the mechanism of unfolding processes.

Original languageEnglish
Pages (from-to)4605-4610
Number of pages6
JournalBiophysical Journal
Volume91
Issue number12
DOIs
Publication statusPublished - Dec 2006

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Temperature
Enzymes
Hot Temperature
Protein Unfolding
Fluorescence
Proteins

ASJC Scopus subject areas

  • Biophysics

Cite this

Enzyme kinetics above denaturation temperature : A temperature-jump/stopped- flow apparatus. / Kintses, Bálint; Simon, Zoltán; Gyimesi, Máté; Tóth, Júlia; Jelinek, Balázs; Niedetzky, Csaba; Kovács, M.; Málnási-Csizmadia, A.

In: Biophysical Journal, Vol. 91, No. 12, 12.2006, p. 4605-4610.

Research output: Contribution to journalArticle

Kintses, Bálint ; Simon, Zoltán ; Gyimesi, Máté ; Tóth, Júlia ; Jelinek, Balázs ; Niedetzky, Csaba ; Kovács, M. ; Málnási-Csizmadia, A. / Enzyme kinetics above denaturation temperature : A temperature-jump/stopped- flow apparatus. In: Biophysical Journal. 2006 ; Vol. 91, No. 12. pp. 4605-4610.
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