Engels Verena, Georgi Tobias, Wendisch Volker F
Institute of Biotechnology I, Research Center Juelich, Juelich, Germany.
FEMS Microbiol Lett. 2008 Dec;289(1):80-9. doi: 10.1111/j.1574-6968.2008.01370.x.
Corynebacterium glutamicum can grow on a variety of carbohydrates from which glucose, fructose and sucrose are taken up and phosphorylated by the phosphoenolpyruvate-dependent phosphotransferase system (PTS). Here, we show that cg2927 (scrB) encodes sucrose-6-phosphate hydrolase. The purified His-tagged protein hydrolyzed sucrose-6-phosphate and sucrose, but not sucrose-6'-phosphate. The Km value for sucrose was 190 mM while the Km for sucrose-6-phosphate was much lower, 0.04 mM. Sucrose-6-phosphate hydrolase activity was stimulated by MgSO4 and fructose-6-phosphate and was inhibited by MnCl2, CaCl2, CuSO4 and ZnSO4. A scrB deletion mutant could not grow on sucrose as the sole carbon source. In addition, growth in the absence of scrB was severely decreased when sucrose was present in addition to glucose, fructose or acetate, suggesting that higher intracellular concentrations of sucrose-6-phosphate are toxic. Transcriptional start sites in the cg2929-cg2928-scrB-ptsS locus could be revealed upstream of cg2929 and upstream of the sucrose-specific PTS gene ptsS. Of these, only ptsS showed increased expression when grown in the presence of sucrose, which was due to control by the transcriptional regulator SugR. The sucrose-6-phosphate hydrolase activity, however, was increased two- to threefold during growth in fructose- or sucrose-containing media, regardless of the presence or absence of SugR.
谷氨酸棒杆菌能够利用多种碳水化合物生长,其中葡萄糖、果糖和蔗糖可通过磷酸烯醇式丙酮酸依赖性磷酸转移酶系统(PTS)被摄取并磷酸化。在此,我们表明cg2927(scrB)编码蔗糖-6-磷酸水解酶。纯化的带有His标签的蛋白质可水解蔗糖-6-磷酸和蔗糖,但不能水解蔗糖-6'-磷酸。蔗糖的Km值为190 mM,而蔗糖-6-磷酸的Km值则低得多,为0.04 mM。蔗糖-6-磷酸水解酶活性受到MgSO4和6-磷酸果糖的刺激,并受到MnCl2、CaCl2、CuSO4和ZnSO4的抑制。scrB缺失突变体不能以蔗糖作为唯一碳源生长。此外,当除葡萄糖、果糖或乙酸盐外还存在蔗糖时,缺乏scrB时的生长会严重下降,这表明细胞内较高浓度的蔗糖-6-磷酸具有毒性。在cg2929-cg2928-scrB-ptsS基因座中,转录起始位点可在cg2929上游和蔗糖特异性PTS基因ptsS上游被揭示。其中,只有ptsS在蔗糖存在下生长时表达增加,这是由于转录调节因子SugR的控制。然而,无论SugR是否存在,在含果糖或蔗糖的培养基中生长期间,蔗糖-6-磷酸水解酶活性都会增加两到三倍。