Inactivation of the ptsI gene encoding enzyme I of the sugar phosphotransferase system of Streptococcus salivarius:: effects on growth and urease expression
Inactivation of the ptsI gene encoding enzyme I of the sugar phosphotransferase system of Streptococcus salivarius:: effects on growth and urease expression
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DOI：
10.1099/00221287-146-5-1179
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发表时间：
2000-05-01
期刊：
影响因子：
2.8
通讯作者：
Burne, RA
中科院分区：
文献类型：
--
作者：
Weaver, CA;Chen, YYM;Burne, RA
唾液链球菌57.1的尿素酶基因在中性ph值下生长的细胞中被紧密抑制。当细胞在酸性ph值下培养时，当细胞在碳水化合物过量的条件下生长时，尿素酶基因被去抑制，转录被增强。此前，作者提出，当碳水化合物有限时，细菌糖：磷酸转移酶系统(PTS)通过磷酸化尿素酶抑制物来调节DNA结合活性。这项研究的目的是评估PTS的酶I(EI)是否参与调节尿素酶的表达，以响应碳水化合物的供应。通过插入失活pts1基因构建了一株缺失EL的唾液链霉菌57.1株(ptsl18-3)。经Western分析，该突变体没有可测量的PTS活性，并且缺乏EL。突变体在非PTS糖乳糖上的生长情况与野生型菌株一样好，当另一种非PTS糖半乳糖为唯一碳水化合物时，突变体的生长情况好于亲本。该突变株能够在葡萄糖为唯一碳水化合物的条件下生长，但表现出24小时的滞后时间，世代时间是菌株57.1的3倍。Ptsl18-3在添加果糖后48h的平均OD600为0.16，3d后仍未观察到额外的生长。在所测试的所有条件下，两株菌株在中性ph值下都能抑制尿素酶。野生型细胞在有限浓度的乳糖上生长，与在PTS糖上生长相比，尿素酶的表达水平非常低。相反，在类似的条件下，ptsl18-3中的尿素酶表达恢复到在PTS糖上生长的亲本的水平。在过量乳糖条件下的生长进一步抑制了尿素酶的表达，但ptsl18-3表达的尿素酶活性是57.1的3倍左右。这些结果支持EL在尿素酶调控中的作用，但也表明额外的因素可能在调节尿素酶基因的表达中起重要作用。
The urease genes of Streptococcus salivarius 57.1 are tightly repressed in cells growing at neutral ph. When cells are cultivated at acidic ph values, the urease genes become derepressed and transcription is enhanced when cells are growing under carbohydrate-excess conditions. Previously, the authors proposed that the bacterial sugar: phosphotransferase system (PTS) modulated the DNA-binding activity by phosphorylation of the urease repressor when carbohydrate was limiting. The purpose of this study was to assess whether enzyme I (EI) of the PTS could be involved in modulating urease expression in response to carbohydrate availability. An El-deficient strain (ptsl18-3) of S. salivarius 57.1 was constructed by insertional inactivation of the ptsl gene. The mutant had no measurable PTS activity and lacked El, as assessed by Western analysis. The mutant grew as well as the wild-type strain on the non-PTS sugar lactose, and grew better than the parent when another non-PTS sugar, galactose, was the sole carbohydrate. The mutant was able to grow with glucose as the sole carbohydrate, but displayed a 24 h lag time and had a generation time some threefold longer than strain 57.1. The mean OD600 attained after 48 h by ptsl18-3 supplied with fructose was 0.16, with no additional growth observed even after 3 d. Urease expression in the wild-type and mutant strains was assessed in continuous chemostat culture. Repression of urease at neutral ph was seen in both strains under all conditions tested. Growth of wild-type cells on limiting concentrations of lactose resulted in very low levels of urease expression compared with growth on PTS sugars. In contrast, under similar conditions, urease expression in ptsl18-3 was restored to levels seen in the parent growing on PTS sugars. Growth under conditions of lactose excess resulted in further derepression of urease, but ptsl18-3 expressed about threefold higher urease activity than 57.1. The results support a role for El in urease regulation, but also indicate that additional factors may be important in regulating urease gene expression.