Purification of an indole alkaloid biosynthetic enzyme, strictosidine synthase, from a recombinant strain of Escherichia coli.
The gene for the indole alkaloid biosynthetic enzyme, strictosidine synthase, of Catharanthus roseus has been cloned into an inducible Escherichia coli expression vector using an expression cassette polymerase chain reaction technique. Induction of the gene resulted in overexpression of the enzyme which accumulated mainly as insoluble inclusion bodies. Denaturation and refolding of the insoluble protein resulted in the ability to purify up to 6 mg of active enzyme from a single liter of cell culture. The recombinant enzyme has good activity (approximately 30 nkat/mg).
Roessner
C A
CA
Chemistry Department, Texas A&M University, College Station 77843-3255.
Devagupta
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R
Hasan
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M
Williams
H J
HJ
Scott
A I
AI
D000470:Alkaloids; D001483:Base Sequence; D019759:Carbon-Nitrogen Lyases; D004926:Escherichia coli / Q000235:genetics; D002479:Inclusion Bodies / Q000201:enzymology; D008969:Molecular Sequence Data; D010946:Plants, Medicinal / Q000201:enzymology / Q000235:genetics*; D014176:Protein Biosynthesis; D011487:Protein Conformation; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D014158:Transcription, Genetic; D014166:Transferases / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification
D000470:Alkaloids; D011994:Recombinant Proteins; D014166:Transferases; D019759:Carbon-Nitrogen Lyases; C022684:strictosidine synthetase