Thermostable tyrosine phenol-lyase of Symbiobacterium sp. SC-1 : gene cloning, sequence determination, and overproduction in Escherichia coli

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Title
Thermostable tyrosine phenol-lyase of Symbiobacterium sp. SC-1 : gene cloning, sequence determination, and overproduction in Escherichia coli
Author(s)
Seung Goo Lee; Seung Pyo Hong; Yoon Ho Choi; Yong Joon Chung; Moon Hee Sung
Bibliographic Citation
Protein Expression and Purification, vol. 11, pp. 263-270
Publication Year
1997
Abstract
During the screening for tyrosine phenol-lyase-producing thermophiles, we isolated an obligatory symbiotic thermophile, Symbiobacterium sp. SC-1, which grew only in coculture with Bacillus sp. SK-1. A gene encoding thermostable tyrosine phenol-lyase (TPL) was cloned from the genomic DNA of the Symbiobacterium sp. SC-1 and the nucleotide sequence of the TPL structural gene was determined. The gene consists of 1374 base pairs encoding a polypeptide of 458 amino acid residues; the molecular mass of the enzyme subunit is estimated to be 52,196 Da. The structural gene of TPL was amplified by PCR, blunt-ended, and ligated into the NcoI-HindIII site of plasmid pTrc99A to construct an expression vector for the overproduction of the thermostable TPL. The level of thermostable TPL production was about 15% of the total soluble proteins of Escherichia coli extract. The enzyme was purified to homogeneity from the E. coli extract with an overall yield of 48%.
ISSN
1046-5928
Publisher
Elsevier
DOI
http://dx.doi.org/10.1006/prep.1997.0792
Type
Article
Appears in Collections:
Synthetic Biology and Bioengineering Research Institute > 1. Journal Articles
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