Guan H P, Baba T, Preiss J
Department of Biochemistry, Michigan State University, East Lansing 48824.
Cell Mol Biol (Noisy-le-grand). 1994 Nov;40(7):981-8.
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.
一个编码玉米分支酶II（BEII）的cDNA克隆已从玉米胚乳cDNA文库中独立分离出来。将玉米BEII的推导蛋白序列与来自不同来源的BE序列进行了比较。利用T7启动子在大肠杆菌中表达了编码玉米胚乳成熟BEII的基因。表达的BEII被纯化至接近均一，从而可以从BE制剂中完全消除淀粉分解活性和细菌BE。表达的酶与从发育中的玉米胚乳中纯化的BEII具有非常相似的特性。这一结果证实了我们早期的报道，即BEII支链直链淀粉的速率较低，支链淀粉的支化速率是支链直链淀粉的两倍。该研究还表明，从细菌表达系统中纯化BEII比从发育中的玉米胚乳中纯化具有更大的优势。最重要的是，该研究建立了一个利用定点诱变研究玉米BE结构-功能关系的有用工具。
