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Title 

Enhanced stability of tyrosine phenol-lyase from Symbiobacterium toebii by DNA shuffling

Authors 

Jin Ho KimJae Jun SongB G KimM H SungSang Chul Lee

Publisher 

The Korean Society for Applied Microbiology

Issue Date 

2004

Citation 

Journal of Microbiology and Biotechnology, vol. 14, no. 1, pp. 153-157

Keywords 

denaturant toleranceDNA shufflingthermostabilitytyrosine phenol-lyaseDNAtyrosine phenol lyaseDNA screeningDNA strandenzyme stability

Abstract 

Tyrosine phenol-lyase (TPL) is a useful enzyme for the synthesis of pharmaceutical aromatic amino acids. In the current study, sequential DNA shuffling and screening were used to enhance the stability of TPL. Twenty-thousand mutants were screened, and several improved variants were isolated. One variant named A13V, in which the 13th amino acid alanine was substituted by valine, exhibited a higher temperature and denaturant stability than the wild-type TPL. The purified mutant TPL, A13V, retained about 60% of its activity at 76°C, whereas the activity of the wild-type TPL decreased to less than 20% at the same temperature. Plus, A13V exhibited about 50% activity with 3 M urea, while the wild-type TPL lost almost all its catalytic activity, indicating an increased denaturant tolerance in the mutant A13V. It is speculated that the substitution of Val for the Ala in the β-strand of the N-terminal arm was responsible for the heightened stabilization, and that the current results will contribute to further research on the structural stability of TPL.

ISSN 

1017-7825

Appears in Collections

1. Journal Articles > Journal Articles

Registered Date

2019-05-02


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