Saffen D W, Presper K A, Doering T L, Roseman S
Department of Biology, Johns Hopkins University, Baltimore, Maryland 21218.
J Biol Chem. 1987 Nov 25;262(33):16241-53.
Specialized lambda-transducing phages that carry the Escherichia coli genes ptsH, ptsI, crr, cysM, and cysA have been isolated, and the genes were subcloned in plasmid pBR322. Subcloning and restriction mapping data gave the following clockwise order of genes located at about 52 min on the E. coli genetic map: lig, cysK, ptsH, ptsI, crr, cysM, cysA. The nucleotide sequences of ptsH, ptsI, and crr and the corresponding flanking regions have been determined. These genes encode three cytoplasmic proteins of the phosphoenol-pyruvate:glycose phosphotransferase system: HPr, Enzyme I, and IIIGlc, respectively. The deduced amino acid sequences are consistent with amino acid composition and Edman degradation analyses obtained with the purified proteins. The calculated subunit molecular weight values (9,109 for HPr, 63,489 for Enzyme I, and 18,099 for IIIGlc) also agree well with values obtained with the proteins. Results of gamma delta-transposon insertional studies provided definitive evidence that IIIGlc is the gene product of crr, and therefore that IIIGlc plays a critical role in regulating the metabolism and uptake of certain non-PTS sugars (see accompanying papers: Mitchell, W.J., Saffen, D.W., and Roseman, S. (1987) J. Biol. Chem. 16254-16260; Misko, T.P., Mitchell, W.J., Meadow, N.D., and Roseman, S. (1987) J. Biol. Chem. 16261-16266). The gamma delta transposon studies also suggest that crr is transcribed from an independent promoter located within the ptsI gene. Putative regulatory sequence features include a catabolite gene activator protein-cAMP-binding site and two regions of 2-fold rotational symmetry adjacent to the potential promoter upstream from the HPr structural gene, several ribosome-binding sites, and a rho-independent RNA polymerase termination site downstream from crr. In addition, the ptsI gene contains two highly conserved direct repeats. The significance of these sequence features is discussed with respect to possible multiple forms of pts regulation.
携带大肠杆菌基因ptsH、ptsI、crr、cysM和cysA的特异性λ转导噬菌体已被分离出来，这些基因被亚克隆到质粒pBR322中。亚克隆和限制性图谱分析数据给出了位于大肠杆菌遗传图谱约52分钟处的基因按顺时针方向的顺序：lig、cysK、ptsH、ptsI、crr、cysM、cysA。已确定了ptsH、ptsI和crr以及相应侧翼区域的核苷酸序列。这些基因分别编码磷酸烯醇丙酮酸：葡萄糖磷酸转移酶系统的三种细胞质蛋白：HPr、酶I和IIIGlc。推导的氨基酸序列与通过纯化蛋白获得的氨基酸组成和埃德曼降解分析结果一致。计算得到的亚基分子量值（HPr为9109，酶I为63489，IIIGlc为18099）也与通过蛋白获得的值非常吻合。γδ转座子插入研究的结果提供了确凿证据，表明IIIGlc是crr的基因产物，因此IIIGlc在调节某些非磷酸转移酶系统（PTS）糖类的代谢和摄取中起关键作用（见随附论文：米切尔，W.J.，萨芬，D.W.，和罗斯曼，S.（1987年）《生物化学杂志》16254 - 16260；米斯科，T.P.，米切尔，W.J.，梅多，N.D.，和罗斯曼，S.（1987年）《生物化学杂志》16261 - 16266）。γδ转座子研究还表明，crr是从位于ptsI基因内的一个独立启动子转录而来的。推测的调控序列特征包括一个分解代谢基因激活蛋白 - cAMP结合位点以及在HPr结构基因上游潜在启动子附近的两个具有2倍旋转对称性的区域、几个核糖体结合位点，以及crr下游的一个不依赖ρ因子的RNA聚合酶终止位点。此外，ptsI基因包含两个高度保守的正向重复序列。关于pts调控可能的多种形式，讨论了这些序列特征的意义。
