Tian G, Lim D, Oppenheim J D, Maas W K
Department of Microbiology, New York University Medical Center, NY 10016.
J Mol Biol. 1994 Jan 7;235(1):221-30. doi: 10.1016/s0022-2836(05)80028-2.
In Escherichia coli K12, formation of the enzymes of arginine biosynthesis are controlled by arginine, with complete repression during growth with added arginine, severe repression (about 95%) during growth without added arginine and complete derepression during arginine-limited growth. In E. coli B, the degree of repression is not correlated with arginine concentrations. Under all conditions of growth enzyme formation is repressed, with repression being somewhat less in a medium with arginine than in a medium without arginine. These differences in repressibility between the two strains have been shown previously to be due to the presence of different alleles of argR, the gene for the arginine repressor. Here we have compared the binding of the two repressors to the operator sites of argF (ARG boxes). In DNase I footprinting and gel retardation experiments with argF ARG boxes we have shown that the arginine repressor of E. coli K12 bound to arginine (ArgRK-arg) has a greater affinity than the arginine repressor of E. coli B bound to arginine (ArgRB-arg), whereas free ArgRB (ArgRBf) has a much stronger affinity than free ArgRK (ArgRKf). The stronger binding of ArgRBf can explain the repression seen in E. coli B during arginine-limited growth and indicates that ArgRBf, but not ArgRKf, is able to repress enzyme synthesis under physiological conditions. The weaker repression of E. coli B than of E. coli K12 seen in the presence of arginine can be explained by the lower affinity of ArgRB-arg for operator sites as compared to ArgRK-arg. Another contributing cause for the weaker repression is the reduction of ArgRBf concentration due to autoregulation of the gene for the repressor. Thus the combined effects of repression by ArgRBf, but not ArgRKf, with the weaker repression by ArgRB-arg as compared to ArgRK-arg, convert the arginine dependent regulation in E. coli K12 to arginine independent regulation in E. coli B.
在大肠杆菌K12中,精氨酸生物合成酶的形成受精氨酸控制,在添加精氨酸的情况下生长时会完全被抑制,在不添加精氨酸的情况下生长时会受到严重抑制(约95%),而在精氨酸限制生长期间则会完全去抑制。在大肠杆菌B中,抑制程度与精氨酸浓度无关。在所有生长条件下,酶的形成都会受到抑制,在含有精氨酸的培养基中,抑制程度比在不含精氨酸的培养基中略低。先前已表明,这两种菌株在可抑制性方面的差异是由于精氨酸阻遏物基因argR存在不同等位基因所致。在此,我们比较了两种阻遏物与argF(ARG框)的操纵位点的结合情况。在使用argF ARG框进行的DNase I足迹实验和凝胶阻滞实验中,我们发现与精氨酸结合的大肠杆菌K12精氨酸阻遏物(ArgRK-arg)比与精氨酸结合的大肠杆菌B精氨酸阻遏物(ArgRB-arg)具有更高的亲和力,而游离的ArgRB(ArgRBf)比游离的ArgRK(ArgRKf)具有更强的亲和力。ArgRBf更强的结合能力可以解释在大肠杆菌B中精氨酸限制生长期间观察到的抑制现象,这表明ArgRBf而非ArgRKf能够在生理条件下抑制酶的合成。在存在精氨酸的情况下,大肠杆菌B比大肠杆菌K12的抑制作用较弱,可以通过与ArgRK-arg相比,ArgRB-arg对操纵位点的亲和力较低来解释。另一个导致抑制作用较弱的原因是由于阻遏物基因的自动调节导致ArgRBf浓度降低。因此,ArgRBf而非ArgRKf的抑制作用与ArgRB-arg相比ArgRK-arg较弱的抑制作用共同作用,将大肠杆菌K12中依赖精氨酸的调节转变为大肠杆菌B中不依赖精氨酸的调节。