The esterase from Alicyclobacillus acidocaldarius as a reporter enzyme and affinity tag for protein biosynthesis.
Esterase from thermophilic bacteria Alicyclobacillus acidocaldarius can be produced up to 200 microg/ml by coupled in vitro transcription/translation system derived from Escherichia coli. The synthesized thermostable enzyme can be determined by photometrical and fluorescent assays at least up to 10(-8) M concentration or by activity staining in the polyacrylamide gels. Enhanced green fluorescence protein-esterase fusion protein was bound to a matrix with immobilized esterase inhibitor and purified by affinity chromatography. Thus, the esterase is suited as a reporter enzyme to monitor the expression of polypeptides coupled to its N-terminus and simultaneously, as a cleavable tag for polypeptide purification.
Agafonov
Dmitry E
DE
Laboratorium für Biochemie, Universität Bayreuth, 95440 Bayreuth, Germany.
Rabe
Kersten S
KS
Grote
Michael
M
Huang
Yiwei
Y
Sprinzl
Mathias
M
D000345:Affinity Labels; D001483:Base Sequence; D017931:DNA Primers; D004591:Electrophoresis, Polyacrylamide Gel; D004950:Esterases / Q000378:metabolism*; D016937:Gram-Positive Endospore-Forming Rods / Q000201:enzymology*; D015202:Protein Engineering
D000345:Affinity Labels; D017931:DNA Primers; D004950:Esterases