Cloning of dextransucrase gene from Leuconostoc citreum HJ-P4 and its high-level expression in E. coli by low temperature induction.
A dextransucrase (LcDS) gene from Leuconostoc citreum HJ-P4 has been amplified and cloned in E. coli. The LcDS gene consists of 4,431 nucleotides encoding 1,477 amino acid residues sharing 63-98% of amino acid sequence identities with other known dextransucrases from Leuc. mesenteroides. Interestingly, 0.1 mM of IPTG induction at 15 degrees remarkably increased the LcDS productivity to 19,187 U/l culture broth, which was over 330-fold higher than that induced at 37 degrees. Optimal reaction temperature and pH of LcDS were determined as 35 degrees and pH 5.5 in 20 mM sodium acetate buffer, respectively. Meanwhile, 0.1 mM CaCl(2) increased its activity to the maximum of 686 U/mg, which was 2.1-fold higher than that in the absence of calcium ion. Similar to the native Leuconostoc dextransucrase, recombinant LcDS could successfully produce a series of isomaltooligosaccharides from sucrose and maltose, on the basis of its transglycosylation activity.
Yi
Ah-Rum
AR
Department of Food Science and Technology, Chungbuk National University, Cheongju 361-763, Korea.
Lee
So-Ra
SR
Jang
Myoung-Uoon
MU
Park
Jung-Mi
JM
Eom
Hyun-Ju
HJ
Han
Nam Soo
NS
Kim
Tae-Jip
TJ
D002122:Calcium Chloride / Q000008:administration & dosage; D003001:Cloning, Molecular; D003080:Cold Temperature; D004789:Enzyme Activation / Q000187:drug effects; D004790:Enzyme Induction / Q000187:drug effects*; D004926:Escherichia coli / Q000378:metabolism*; D005798:Genes, Bacterial; D005964:Glucosyltransferases / Q000096:biosynthesis* / Q000235:genetics / Q000302:isolation & purification; D006863:Hydrogen-Ion Concentration; D007218:Industrial Microbiology; D007544:Isopropyl Thiogalactoside / Q000008:administration & dosage; D007934:Leuconostoc / Q000201:enzymology* / Q000235:genetics; D011994:Recombinant Proteins / Q000378:metabolism
D011994:Recombinant Proteins; D007544:Isopropyl Thiogalactoside; D005964:Glucosyltransferases; C020025:dextransucrase; D002122:Calcium Chloride