苯丙氨酸合成的关键酶基因aroG与pheA串联表达

aroG和pheA是与苯丙氨酸合成有关的两个重要基因。在大肠杆菌(Escherichia coli)中，aroG基因编码脱氧阿拉伯糖型庚酮糖磷酸合酶(DS)，该酶催化由糖代谢中心途径分流出来的磷酸烯醇丙酮酸(PEP)和赤鲜糖4磷酸(E4P)缩合形成脱氧阿拉伯糖型庚酮糖磷酸(DAHP)的反应；pheA基因编码一个双功能酶蛋白，它同时催化两步关键反应，即具有分枝酸变位酶(CM)和预苯酸脱水(PD)的两种功能。采用PCR技术分别从两个不同品系的大肠杆菌染色体DNA中扩增到aroG和pheA。当这两个基因串联在一个质粒上导入大肠杆菌P2392中进行表达时，它们编码的酶DS、CM和PD活性分别提高43、44和22倍；导入短杆菌(Brevibacterium)2731中表达时，相应的酶活性分别提高123、23和56倍。两基因的串联表达能大幅度地提高工程菌株的苯丙氨酸发酵产量。

EXPRESSION OF GENES aroG AND pheA IN PHENYLALANINE BIOSYNTHESIS

aroGand pheAgenes,encoding 3 Deoxy D arabinoheptulonate 7 phosphatesyn thase(DS) and Chorismate mutase (CM) prephenate dehydratase(PD) in the pathway of phenylalnine biosynthesisrespectively,were amplified by polymerase chain reaction(PCR) .The genes were assembled on the multicopy vectors and expressed in both Escherichia coli and Brevibacterium .The productsoftwogene weredetected by SDS PAGE.Theactivities ofrelevantenzymes were measuredinthecrudeextractofthe hoststrain. When aroG pheA genes wereintroducedinto E.coli p2392 ,the activities of DS,CMand PDwereincreased by 4 .3 fold,4 .4 fold and 2.2 fold respectively. Whereas in the case of Brevibacterium flavum 2732,the activitiesof DS,CMand PDwereincreased by 12.3 fold,2.3 fold and 5 .6 fold,respectively. As the results,the overproduction of phenylalanine was brought about by using the geneticengineeringstrain of B.flavum .