Biosynthesis of active spinach-chloroplast thioredoxin f in transformed E. coli.
The recently cloned gene for spinach chloroplast thioredoxin f was subcloned in a modified pKK233-2 expression vector and used for transformation of Escherichia coli cells containing the Iq plasmid. After induction with IPTG (isopropyl-beta-D-thiogalactoside) the transformed cells produce the chloroplast protein in large amounts as insoluble deposit within the cell. The protein has been solubilized, purified and analysed for activity. It shows no difference in catalytic activity from native spinach chloroplast thioredoxin f. Its electrophoretic behaviour suggests that the native thioredoxin f may have a different N-terminus than was assumed on the basis of the protein sequencing results.
Aguilar
F
F
Laboratoire de Biochimie végétale, Université de Neuchâtel, Switzerland.
Brunner
B
B
Gardet-Salvi
L
L
Stutz
E
E
Schürmann
P
P
D000595:Amino Acid Sequence; D001483:Base Sequence; D054479:Chloroplast Thioredoxins; D002736:Chloroplasts / Q000378:metabolism*; D003001:Cloning, Molecular; D004247:DNA; D004926:Escherichia coli / Q000235:genetics*; D015151:Immunoblotting; D007525:Isoelectric Focusing; D008969:Molecular Sequence Data; D010940:Plant Proteins / Q000096:biosynthesis / Q000235:genetics* / Q000302:isolation & purification; D010944:Plants / Q000235:genetics*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification; D013879:Thioredoxins; D014169:Transformation, Bacterial
D054479:Chloroplast Thioredoxins; D010940:Plant Proteins; D011994:Recombinant Proteins; D013879:Thioredoxins; D004247:DNA