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Over-expression, purification, and characterization of recombinant NAD-malic enzyme from Escherichia coli K12.

Abstract：

:NAD(+)-dependent malic enzyme (NAD-ME) gene from Escherichia coli K12 was inserted into an expression vector pET24b(+) and transformed into E. coli BL21 (DE3). Recombinant NAD-ME was expressed upon IPTG induction, purified with affinity chromatography, and biochemically characterized. The results showed that recombinant NAD-ME could be produced mainly in a soluble form. The monomeric molecular weight of recombinant NAD-ME was about 65 kDa, whereas monomer, homotetramer, and homooctamer were formed in solution as revealed by nondenaturing polyacrylamide gel electrophoresis analysis. Finally, the K(m) values of NAD-ME for L-malate and NAD were determined as 0.420+/-0.174 and 0.097+/-0.038 mM, respectively, at pH 7.2. By using this over-expression and purification system, recombinant E. coli K12 NAD-ME can now be obtained in large quantity necessary for further biochemical characterization and applications.

journal_name

Protein Expr Purif

journal_title

Protein expression and purification

authors

Wang J,Tan H,Zhao ZK

doi

10.1016/j.pep.2006.11.017

subject

Has Abstract

pub_date

2007-05-01 00:00:00

pages

97-103

issue

1

eissn

1046-5928

issn

1096-0279

pii

S1046-5928(06)00393-7

journal_volume

53

pub_type

杂志文章

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