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Overproduction and crystallization of tryptophanase from recombinant cells of Escherichia coli
Author(s) -
Tani S.,
Tsujimoto N.,
Kawata Y.,
Tokushige M.
Publication year - 1990
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1111/j.1470-8744.1990.tb00078.x
Subject(s) - tryptophanase , escherichia coli , pbr322 , enzyme , polyethylene glycol , plasmid , overproduction , chemistry , recombinant dna , biochemistry , potassium phosphate , microbiology and biotechnology , biology , gene , chromatography
We have cloned the tryptophanase structural gene from Escherichia coli B/1t7‐A into E. coli K‐12 MD55 with a vector plasmid, pBR322. The cloned cells produced a large amount of the enzyme corresponding to more than 30% of the total soluble protein. With the enzyme obtained by this overproduction system, we have prepared three different crystals of tryptophanase, apo‐enzyme, holo‐enzyme, and a complex of holo‐enzyme and L‐alanine, by using polyethylene glycol 4000 or potassium phosphate as a precipitant and the hanging drop method. These single crystals appeared to be suitable for X‐ray diffraction analysis.