Expression, Purification, and Characterization of the Recombinant NAD-Malic Enzyme fromAscaris suum

The cDNA encoding the 65-kDa subunit of malic enzyme fromAscaris suumwas cloned into the bacterial expression vector pKK223-3 and overproduced inEscherichia coli.A protein with a subunit molecular mass of 65,000 was expressed at a level of up to 3% of the total soluble protein in JM109, as judged by...

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Bibliographic Details
Published in:Protein expression and purification 1997-06, Vol.10 (1), p.51-54
Main Authors: Chooback, Lilian, Karsten, William E., Kulkarni, Gopal, Nalabolu, Srinivasa R., Harris, Ben G., Cook, Paul F.
Format: Article
Language:English
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Summary:The cDNA encoding the 65-kDa subunit of malic enzyme fromAscaris suumwas cloned into the bacterial expression vector pKK223-3 and overproduced inEscherichia coli.A protein with a subunit molecular mass of 65,000 was expressed at a level of up to 3% of the total soluble protein in JM109, as judged by SDS–PAGE. The enzyme was purified using column chromatography on phenyl-Sepharose followed by orange-A agarose. The purification procedure resulted in a 32-fold purification with an overall yield of 51%. The bacterially expressed enzyme exhibits kinetic constants identical to those measured for nativeA. suumNAD-malic enzyme.
ISSN:1046-5928
1096-0279
DOI:10.1006/prep.1996.0705