PHYSIOLOGICAL CONSEQUENCES OF THE COMPLETE LOSS OF PHOSPHORYL-TRANSFER PROTEINS HPR AND FPR OF THE PHOSPHOENOLPYRUVATE - SUGAR PHOSPHOTRANSFERASE SYSTEM AND ANALYSIS OF FRUCTOSE (FRU) OPERON EXPRESSION IN SALMONELLA-TYPHIMURIUM
PHYSIOLOGICAL CONSEQUENCES OF THE COMPLETE LOSS OF PHOSPHORYL-TRANSFER PROTEINS HPR AND FPR OF THE PHOSPHOENOLPYRUVATE - SUGAR PHOSPHOTRANSFERASE SYSTEM AND ANALYSIS OF FRUCTOSE (FRU) OPERON EXPRESSION IN SALMONELLA-TYPHIMURIUM
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DOI：
10.1128/jb.172.9.5459-5469.1990
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发表时间：
1990-09-01
影响因子：
3.2
通讯作者：
SAIER, MH
中科院分区：
文献类型：
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作者：
FELDHEIM, DA;CHIN, AM;SAIER, MH
突变体的鼠伤寒沙门氏菌缺陷的果糖操纵子[fruB（MH）KA]，果糖阻遏物（fruR），能量偶联酶的磷酸烯醇丙酮酸：糖磷酸转移酶系统（PTS）（ptsH和ptsI），和蛋白质的环磷酸腺苷行动（cya和crp）的细胞生理过程和果糖操纵子的表达进行了分析。fru操纵子由三个结构基因组成：fruB（MH），其编码PTS的酶IIIFru-调节剂-FPr三结构域融合蛋白; fruK，其编码果糖-1-磷酸激酶;和fruA，其编码PTS的酶IIFru。其中分析的突变体是Tn10插入突变体和lacZ转录融合突变体。发现fruR：：Tn10插入突变体、几种fruB（MH）：：Mu dJ和fruK：：Mu dJ融合突变体和几种ptsH1缺失突变体表达fru操纵子和β-在高组成型水平下，ptsH点突变体和fruA：：Mu dJ融合突变体保留了诱导性。反式包含野生型fru操纵子不能恢复果糖诱导的β-图1示出了在fru：：Mu dJ融合突变体中的半乳糖苷酶表达。CyA和CRP突变体显示所有FRU调节子酶的基础活性降低，但诱导活性未受损。令人惊讶的是，fruB：：Mu dJ crp或cya双突变体显示出超过10倍的抑制的β-在添加果糖后，半乳糖苷酶活性降低，即使这种活性在含有野生型cya和crp等位基因的fruB：Mu dJ融合突变体中仅是轻微可诱导的。相比之下，fruK：：Mu dJ融合突变体中的半乳糖苷酶活性（其通过引入crp或cya突变而被类似地抑制）保持组成型。其他实验表明，通过PTS的糖摄取可以利用FPr-P或HPr-P作为磷酰基供体，但是FPr被称为果糖摄取，而HPr优选用于其他糖的摄取。 缺乏这两种蛋白质的双突变体的所有糖底物的thr PTS的利用率为负，是负的几个heterogeneic碳源的利用率，表现出大大降低腺苷酸环化酶活性，并在很大程度上nonmotile。这些表型特性比紧密ptsH和ptsI突变体（包括这些基因缺失的突变体）所观察到的更为极端。一个生物化学的解释，这一事实提出。
Mutants of Salmonella typhimurium defective in the proteins of the fructose operon [fruB(MH)KA], the fructose repressor (fruR), the energy-coupling enzymes of the phosphoenolpyruvate:sugar phosphotransferase system (PTS) (ptsH and ptsI), and the proteins of cyclic AMP action (cya and crp) were analyzed for their effects on cellular physiological processes and expression of the fructose operon. The fru operon consists of three structural genes: fruB(MH), which encodes the enzyme IIIFru-modulator-FPr tridomain fusion protein of the PTS; fruK, which encodes fructose-1-phosphate kinase; and fruA, which encodes enzyme IIFru of the PTS. Among the mutants analyzed were Tn10 insertion mutants and lacZ transcriptional fusion mutants. It was found that whereas a fruR::Tn10 insertion mutant, several fruB (MH)::Mu dJ and fruK::Mu dJ fusion mutants, and several ptsH1 deletion mutants expressed the fru operon and .beta.-galactosidase at high constitutive levels, ptsH point mutants and fruA::Mu dJ fusion mutants retained inducibility. Inclusion of the wild-type fru operon in trans did not restore fructose-inducuble .beta.-galactosidase expression in the fru::Mu dJ fusion mutants. cya and crp mutants exhibited reduced basal activities of all fru regulon enzymes, but inducibility was not impaired. Surprisingly, fruB::Mu dJ crp or cya double mutants showed over 10-fold inducibility of the depressed .beta.-galactosidase activity upon addition of fructose, even though this activity in the fruB:Mu dJ fusion mutants that contained the wild-type cya and crp alleles was only slightly inducible. By contrast, .beta.-galactosidase activity in a fruK::Mu dJ fusion mutant, which was similarly depressed by introduction of a crp or cya mutation, remained constitutive. Other experiments indicated that sugar uptake via the PTS can utilize either FPr-P or HPr-P as the phosphoryl donor, but that FPr is referred for fructose uptake whereas HPr is preferred for uptake of the other sugars. Double mutants lacking both proteins were negative for the utilization of all sugar substrates of thr PTS, were negative for the utilization of several gluconeogenic carbon sources, exhibited greatly reduced adenylate cyclase activity, and were largely nonmotile. These phenotypic properties are more extreme than those observed for tight ptsH and ptsI mutants, including mutants deleted for these genes. A biochemical explanation for this fact is proposed.