Mitchell W J, Saffen D W, Roseman S
McCollum-Pratt Institute, Johns Hopkins University, Baltimore, Maryland 21218.
J Biol Chem. 1987 Nov 25;262(33):16254-60.
Escherichia coli and Salmonella typhimurium preferentially utilize sugar substrates of the phosphoenol-pyruvate:glycose phosphotransferase system (PTS) when the growth medium also contains other sugars. This phenomenon, diauxic growth, is regulated by the crr gene, which encodes the PTS protein IIIGlc (Saffen, D.W., Presper, K.A., Doering, T.L., and Roseman, S. (1987) J. Biol. Chem. 16241-16253). We have proposed that non-PTS permeases are regulated by their interaction with IIIGlc, and in vitro studies from other laboratories have provided support for this model, but the in vivo effects of excess IIIGlc are not known. In the present studies, transformed cells that overproduced IIIGlc 2- and 10-fold, respectively, were constructed from a pts+ strain of E. coli and plasmids containing the crr gene. In the 2-fold overproducer, fermentation of, and growth on the non-PTS carbohydrates glycerol, lactose, maltose, and melibiose was generally more sensitive to the glucose analogue methyl-alpha-D-glucopyranoside than in a control strain containing normal levels of IIIGlc. In addition, inhibition of lactose permease activity by methyl-alpha-glucoside (inducer exclusion) was more effective in the 2-fold overproducer than in the control strain, particularly when the permease activity was high. The 10-fold IIIGlc overproducing strain had a requirement for the amino acids methionine, isoleucine, leucine, and valine that may or may not be related to the increased concentration of IIIGlc. Fermentation of non-PTS carbohydrates was also poor in the latter strain. Finally, lactose permease activity was 50% of that in control cells containing the same levels of beta-galactosidase, and the lactose permease activity in the IIIGlc overproducer was reduced to an extremely low level in the presence of methyl alpha-glucoside. Thus there is an inverse relationship between the cellular concentration of IIIGlc and the ability to metabolize non-PTS substrates. The results are consistent with the model where inducer exclusion is affected by a direct interaction between IIIGlc and a non-PTS transport system.
当生长培养基中还含有其他糖类时，大肠杆菌和鼠伤寒沙门氏菌优先利用磷酸烯醇式丙酮酸：葡萄糖磷酸转移酶系统（PTS）的糖类底物。这种现象，即双相生长，受crr基因调控，该基因编码PTS蛋白IIIGlc（萨芬，D.W.，普雷斯珀，K.A.，多林，T.L.，和罗斯曼，S.（1987年）《生物化学杂志》16241 - 16253页）。我们提出非PTS通透酶通过与IIIGlc的相互作用受到调控，其他实验室的体外研究为该模型提供了支持，但过量IIIGlc的体内效应尚不清楚。在本研究中，分别从大肠杆菌的pts + 菌株和含有crr基因的质粒构建了过量表达IIIGlc 2倍和10倍的转化细胞。在过量表达2倍的细胞中，与含有正常水平IIIGlc的对照菌株相比，非PTS碳水化合物甘油、乳糖、麦芽糖和蜜二糖的发酵及在其上的生长通常对葡萄糖类似物α - D - 甲基吡喃葡萄糖苷更敏感。此外，α - 甲基葡萄糖苷（诱导物排除）对乳糖通透酶活性的抑制在过量表达2倍的细胞中比在对照菌株中更有效，特别是当通透酶活性较高时。过量表达IIIGlc 10倍的菌株对甲硫氨酸、异亮氨酸、亮氨酸和缬氨酸有需求，这可能与IIIGlc浓度增加有关，也可能无关。后一种菌株中，非PTS碳水化合物的发酵也很差。最后，乳糖通透酶活性是含有相同水平β - 半乳糖苷酶的对照细胞的50%，并且在过量表达IIIGlc的细胞中，α - 甲基葡萄糖苷存在时乳糖通透酶活性降低到极低水平。因此，IIIGlc的细胞浓度与代谢非PTS底物的能力之间存在反比关系。结果与诱导物排除受IIIGlc与非PTS转运系统直接相互作用影响的模型一致。
