Functional expression of human glucose-6-phosphate dehydrogenase in Escherichia coli.
The complete coding sequence for human glucose-6-phosphate-dehydrogenase (G6PD) was inserted downstream from the tac promoter of a plasmid, pJF118EH, which also carries the lacIq repressor gene. When Escherichia coli strains (that are unable to grow on glucose due to the absence of functional zwf (G6PD-) and pgi genes) were transformed with this plasmid (pAC1), they were able to grow on glucose as sole carbon source. The rate of growth on glucose was faster in the presence of the inducer of the tac promoter, isopropyl-beta-D-thiogalactopyranoside (IPTG). Extracts of the transformed cells contained a G6PD activity that was not detectable in the parental strains and that was inducible by IPTG. The G6PD activities from normal E. coli and from pAC1-transformed cells comigrated with human G6PD when subjected to electrophoresis on agarose gels. However, when denatured, the G6PD produced by pAC1 was, like the human enzyme, distinguishable from the E. coli-encoded enzyme on the basis of its immunoreactivity with antibody specific for human G6PD. Therefore, human G6PD can be expressed in E. coli and can function to complement the bacterial enzyme deficiency.
Persico
M G
MG
International Institute of Genetics and Biophysics, Naples, Italy.
Ciccodicola
A
A
Martini
G
G
Rosner
J L
JL
D001483:Base Sequence; D003001:Cloning, Molecular; D004247:DNA / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D005947:Glucose / Q000378:metabolism; D005954:Glucosephosphate Dehydrogenase / Q000096:biosynthesis* / Q000235:genetics / Q000276:immunology; D006801:Humans; D007544:Isopropyl Thiogalactoside / Q000494:pharmacology; D008969:Molecular Sequence Data; D010957:Plasmids; D011401:Promoter Regions, Genetic / Q000187:drug effects; D012097:Repressor Proteins / Q000235:genetics; D014170:Transformation, Genetic
D012097:Repressor Proteins; D007544:Isopropyl Thiogalactoside; D004247:DNA; D005954:Glucosephosphate Dehydrogenase; D005947:Glucose