Huang A M, Lee J I, King S C, Wilson T H
Department of Cellular and Molecular Physiology, Harvard Medical School, Boston, Massachusetts 02215.
J Bacteriol. 1992 Aug;174(16):5436-41. doi: 10.1128/jb.174.16.5436-5441.1992.
Five lacY mutants with amber stop codons at known positions were each placed into 12 different suppressor strains. The 60 amino acid substitutions obtained in this manner were tested for growth on lactose-minimal medium plates and for transport of lactose, melibiose, and thiomethylgalactoside. Most of the amino acid substitutions in the regions of the putative loops (between transmembrane alpha helices) resulted in a reasonable growth rate on lactose with moderate-to-good transport activity. In one strain (glycine substituted for Trp-10), abnormal sugar recognition was found. The substitution of proline for Trp-33 (in the region of the first alpha helix) showed no activity, while four additional substitutions (lysine, leucine, cysteine, and glutamic acid) showed low activity. Altered sugar specificity was observed when Trp-33 was replaced by serine, glutamine, tyrosine, alanine, histidine, or phenylalanine. It is concluded that Trp-33 may be involved directly or indirectly in sugar recognition.
五个在已知位置带有琥珀色终止密码子的乳糖通透酶（lacY）突变体分别被置于12种不同的抑制菌株中。通过这种方式获得的60个氨基酸替换在乳糖基本培养基平板上进行生长测试，并检测其对乳糖、蜜二糖和硫代甲基半乳糖苷的转运情况。在假定的环区域（跨膜α螺旋之间）的大多数氨基酸替换导致在乳糖上有合理的生长速率，并具有中度至良好的转运活性。在一个菌株中（色氨酸-10被甘氨酸取代），发现了异常的糖识别。脯氨酸替换色氨酸-33（在第一个α螺旋区域）没有活性，而另外四个替换（赖氨酸、亮氨酸、半胱氨酸和谷氨酸）活性较低。当色氨酸-33被丝氨酸、谷氨酰胺、酪氨酸、丙氨酸、组氨酸或苯丙氨酸取代时，观察到糖特异性发生改变。得出的结论是，色氨酸-33可能直接或间接参与糖识别。
