Characterization of transmembrane domains 6, 7, and 8 of MalF by mutational analysis
Characterization of transmembrane domains 6, 7, and 8 of MalF by mutational analysis
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DOI：
10.1128/jb.178.8.2255-2262.1996
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发表时间：
1996-04-01
影响因子：
3.2
通讯作者：
Ehrmann, M
中科院分区：
文献类型：
--
作者：
Ehrle, R;Pick, C;Ehrmann, M
利用寡核苷酸诱变分离MalF跨膜片段6、7和8中的突变。MalF是大肠杆菌中结合蛋白依赖的麦芽糖转运系统的细胞质膜组分。目前的结构模型预测了MalF的八个跨膜结构域，MalF侧翼的跨膜区段6、7和8或者是存在于细菌ABC转运蛋白超家族成员的细胞质膜亚基中的EAA-X（3）-G-X（9）-I-X-LP共有区的一部分。分离了具有关于麦芽糖转运系统的底物特异性的两种新表型的突变，一种突变体在最小麦芽糖培养基上生长，但在含有麦芽七糖或麦芽七糖加麦芽糖的培养基上不生长，因此被称为糊精显性阴性。另一类突变导致麦芽糖减但麦芽七糖加表型。九个孤立的突变，导致底物特异性的变化紧密聚集在一个面上的假设跨膜螺旋6。在跨膜结构域7中检测到类似的突变聚类。跨膜片段7中的大多数突变导致蛋白酶敏感或条件表型，相对于MalF功能或两者。跨膜结构域8的突变分布较为随机，大多数跨膜结构域8的突变导致Mal（+）Dex（-）表型，6个Mal（+）抑制突变与跨膜结构域7的2个突变分离，改变了跨膜结构域6或8的氨基酸残基。
Oligonucleotide mutagenesis was used to isolate mutations in membrane-spanning segments 6, 7, and 8 of MalF, MalF is a cytoplasmic membrane component of the binding protein-dependent maltose transport system in Escherichia coli. The current structural model predicts eight transmembrane domains for MalF, Membrane-spanning segments 6, 7, and 8 of MalF flank or are part of the EAA-X(3)-G-X(9)-I-X-LP consensus region present in the cytoplasmic membrane subunits of the bacterial ABC transporter superfamily members, Mutations with two novel phenotypes with respect to substrate specificity of the maltose transport system were isolated, One mutant grew on minimal maltose media but not on media containing either maltoheptaose or maltoheptaose plus maltose and was thus termed dextrin dominant negative. The other class of mutations led to a maltose minus but maltoheptaose plus phenotype. Nine of the isolated mutations leading to changes in substrate specificity were tightly clustered on one face of the postulated transmembrane helix 6. A similar clustering of mutations was detected in transmembrane domain 7. The majority of mutations in membrane-spanning segment 7 led to a protease-sensitive or a conditional phenotype with respect to MalF function or both. Mutations in transmembrane domain 8 appeared to be more randomly distributed, The majority of mutations in membrane-spanning segment 8 caused a Mal(+) Dex(-) phenotype, Six Mal(+) suppressor mutations isolated to two mutations in transmembrane domain 7 changed amino acid residues in membrane-spanning segment 6 or 8.