Prestidge R L, Grandison P M, Chuk D W, Booth R J, Watson J D
Genesis Research and Development Corporation Ltd, Auckland, New Zealand.
Gene. 1995 Oct 16;164(1):129-32. doi: 10.1016/0378-1119(95)00470-q.
The 19-kDa antigen (19Ag) of Mycobacterium tuberculosis (Mt) is a lipoprotein which is released from the organism during growth. In order to study the possible involvement of this antigen in the host protective response against Mt infection, it would be helpful to obtain high-level production of 19Ag from a recombinant organism. We have found that overexpression of the native 19Ag gene in Escherichia coli or yeast leads to products which are aggregated and insoluble. By site-directed mutagenesis of the 19Ag lipoprotein leader sequence, we have generated a mutant gene which directs the production of 19Ag into the periplasmic space of E. coli, from where it can be easily purified in high yield. 19Ag obtained from this mutant construct lacks the lipid-modified N-terminal Cys residue found in the native 19Ag, and is not glycosylated, but is otherwise indistinguishable from 19Ag isolated from Mt culture supernatant.
结核分枝杆菌（Mt）的19 kDa抗原（19Ag）是一种脂蛋白，在细菌生长过程中从菌体释放出来。为了研究该抗原在宿主针对Mt感染的保护性反应中可能发挥的作用，从重组生物体中高水平生产19Ag将有所帮助。我们发现，在大肠杆菌或酵母中天然19Ag基因的过表达会导致产物聚集且不溶。通过对19Ag脂蛋白前导序列进行定点诱变，我们产生了一个突变基因，该基因可将19Ag导向大肠杆菌的周质空间，从那里可以很容易地高产量纯化。从这种突变构建体获得的19Ag缺乏天然19Ag中发现的脂质修饰的N端半胱氨酸残基，且未进行糖基化，但在其他方面与从Mt培养上清液中分离的19Ag没有区别。
