Expression of human tyrosine hydroxylase type I in Escherichia coli as a protease-cleavable fusion protein. Short communication.
Wild-type and N-terminal 35-, 38-, and 44-amino acid-deleted mutants of human tyrosine hydroxylase type 1 (hTH1) fused to maltose-binding protein via the target sequence for a restriction protease were expressed in Escherichia coli and purified. The fused protein was treated with the restriction protease factor Xa or enterokinase to isolate hTH1 from the fused form. The treatment of fused wild-type and 35-amino acid-deleted mutant with factor Xa and enterokinase caused non-specific cleavages in the vicinity of the phosphorylation sites, Ser19 and Ser40, due to the flexible conformation of the N-terminus of hTH1.
Nakashima
A
A
Department of Physiology, School of Medicine, Fujita Health University, Toyoake, Aichi, Japan.
Mori
K
K
Nagatsu
T
T
Ota
A
A
D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics* / Q000378:metabolism; D004765:Enteropeptidase / Q000235:genetics* / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics*; D029968:Escherichia coli Proteins; D006801:Humans; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism; D014446:Tyrosine 3-Monooxygenase / Q000235:genetics / Q000378:metabolism*
D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D002352:Carrier Proteins; D029968:Escherichia coli Proteins; D058619:Maltose-Binding Proteins; D009004:Monosaccharide Transport Proteins; D011993:Recombinant Fusion Proteins; C461599:maltose transport system, E coli; D014446:Tyrosine 3-Monooxygenase; D004765:Enteropeptidase