CRCU

Chemical Research in Chinese Universities ›› 2004, Vol. 20 ›› Issue (1): 50-54.

• Articles • Previous Articles     Next Articles

Directed Evolution of L-Aspartase by Mobility of Domains

GOU Xiao-jun1, LI Shuang2, KONG Xiang-duo2, LIU Wei1, SUN Yan-hong2, ZHANG Jin2   

  1. 1. The Joint Laboratory of Molecular Evolution and Engineering, Department of Bioengineering, Chengdu University, Chengdu 610106, P. R. China;
    2. Key Laboratory for Molecular Enzymology and Engineering of Ministry of Education, Jilin University, Changchun 130023, P. R. China
  • Received:2003-01-09 Online:2004-02-24 Published:2011-08-06

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Abstract: To enhance the relative movement of domains, we inserted a random sequence of fifteen-peptide into the three domains of L-aspartase. By means of directed screening, the three isoforms of monomeric, dimmeric and tetrameric enzymes were obtained. Compared to the wild-type tetrameric L-asparease, these mutants remained 19.7%, 42.3%, and 92% of the enzyme activity, respectively. Moreover, the examination of enzyme properties revealed that their kcat and KM changed in varying degrees, and the optimum pH shifted towards acidic pH, while the dependence of the activity of enzyme on Mg2+ concentration and thermostability increased. Therefore this strategy provides a novel approach to directed evolution of enzymes.

Key words: Directed evolution, Domain, L-aspartase, Mobility