Engineering the thermostability of a TIM-barrel enzyme by rational family shuffling

Szilárd Kamondi, András Szilágyi, László Barna, P. Závodszky

Research output: Contribution to journalArticle

13 Citations (Scopus)

Abstract

A possible approach to generate enzymes with an engineered temperature optimum is to create chimeras of homologous enzymes with different temperature optima. We tested this approach using two family-10 xylanases from Thermotoga maritima: the thermophilic xylanase A catalytic domain (TmxAcat, Topt = 68 °C), and the hyperthermophilic xylanase B (TmxB, Topt = 102 °C). Twenty-one different chimeric constructs were created by mimicking family shuffling in a rational manner. The measured temperature optima of the 16 enzymatically active chimeras do not monotonically increase with the percentage of residues coming from TmxB. Only four chimeras had a higher temperature optimum than TmxAcat, the most stable variant (Topt = 80 °C) being the one in which both terminal segments came from TmxB. Further analysis suggests that the interaction between the N- and C-terminal segments has a disproportionately high contribution to the overall thermostability. The results may be generalizable to other enzymes where the N- and C-termini are in contact.

Original languageEnglish
Pages (from-to)725-730
Number of pages6
JournalBiochemical and Biophysical Research Communications
Volume374
Issue number4
DOIs
Publication statusPublished - Oct 3 2008

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Endo-1,4-beta Xylanases
Temperature
Enzymes
Thermotoga maritima
Catalytic Domain

Keywords

  • (β/α)-Barrel fold
  • Chimera
  • Rational family shuffling
  • Thermal stability
  • Thermotoga maritima
  • Xylanase

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Engineering the thermostability of a TIM-barrel enzyme by rational family shuffling. / Kamondi, Szilárd; Szilágyi, András; Barna, László; Závodszky, P.

In: Biochemical and Biophysical Research Communications, Vol. 374, No. 4, 03.10.2008, p. 725-730.

Research output: Contribution to journalArticle

Kamondi, Szilárd ; Szilágyi, András ; Barna, László ; Závodszky, P. / Engineering the thermostability of a TIM-barrel enzyme by rational family shuffling. In: Biochemical and Biophysical Research Communications. 2008 ; Vol. 374, No. 4. pp. 725-730.
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