A plant metallothionein produced in E. coli.
A metallothionein cDNA was generated from pea (Pisum sativum L.) roots, amplified by PCR and inserted into a plasmid for expression in E. coli. Purification of the resultant product generated 3 pools of cadmium-containing material after DEAE-cellulose chromatography. The amino acid composition of each was in excellent agreement with that predicted for pea metallothionein. A cadmium content of approximately 6 g.atoms per mole of protein was estimated. N-terminal sequence analysis revealed that the recombinant molecule had been proteolysed within the extended region linking the 2 cysteine-rich (putative) metal-binding regions. The significance of these findings in terms of the protein folding/targeting of the molecule are considered.
Kille
P
P
Department of Biochemistry, University of Wales College of Cardiff, UK.
Winge
D R
DR
Harwood
J L
JL
Kay
J
J
D000595:Amino Acid Sequence; D001483:Base Sequence; D002850:Chromatography, Gel; D002851:Chromatography, High Pressure Liquid; D002852:Chromatography, Ion Exchange; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics / Q000302:isolation & purification; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D007887:Fabaceae / Q000235:genetics*; D008668:Metallothionein / Q000235:genetics* / Q000302:isolation & purification; D008969:Molecular Sequence Data; D010946:Plants, Medicinal; D010957:Plasmids; D016133:Polymerase Chain Reaction / Q000379:methods; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000302:isolation & purification
D011994:Recombinant Proteins; D004247:DNA; D008668:Metallothionein