Expression of branching enzyme II of maize endosperm in Escherichia coli.
A cDNA clone encoding maize branching enzyme II (BEII) has been independently isolated from a maize endosperm cDNA library. The deduced protein sequence of maize BEII was compared with that of BE from diverse sources. The gene encoding mature BEII of maize endosperm has been expressed in E. coli using the T7 promoter. The expressed BEII was purified to near homogeneity so that amylolytic activity and bacterial BE could be completely eliminated from the BE preparation. The expressed enzyme showed very similar properties to those of BEII purified from developing maize endosperm. This result confirmed our earlier report that BEII had a lower rate of branching amylose and the rate of branching amylopectin was twice that of branching amylose. This study also showed a greater advantage of purifying BEII from the bacterial expression system than from developing maize endosperm. Most importantly, this study has established a useful tool to study the structure-function relationships of the maize BE using site-directed mutagenesis.
Department of Biochemistry, Michigan State University, East Lansing 48824.
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D015061:1,4-alpha-Glucan Branching Enzyme / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D018076:DNA, Complementary / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D017343:Genes, Plant; D006801:Humans; D007527:Isoenzymes / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D008969:Molecular Sequence Data; D017386:Sequence Homology, Amino Acid; D013379:Substrate Specificity; D003313:Zea mays / Q000201:enzymology* / Q000235:genetics*
D017931:DNA Primers; D018076:DNA, Complementary; D007527:Isoenzymes; D015061:1,4-alpha-Glucan Branching Enzyme