Drexler K, Dannull J, Hindennach I, Mutschler B, Henning U
Max-Planck-Institut für Biologie, Tübingen, Germany.
J Mol Biol. 1991 Jun 20;219(4):655-63. doi: 10.1016/0022-2836(91)90662-p.
The T-even type Escherichia coli phage Ox2 recognizes the outer membrane protein OmpA as a receptor. This recognition is accomplished by the 266 residue protein 38, which is located at the free ends of the virion's long tail fibers. Host-range mutants had been isolated in three consecutive steps: Ox2----Ox2h5----Ox2h10----Ox2h12, with Ox2h12 recognizing the outer membrane protein OmpC efficiently and having lost some affinity for OmpA. Protein 38 consists, in comparison with these proteins of other phages, of two constant and one contiguous array of four hypervariable regions; the alterations leading to Ox2h12 were all found within the latter area. Starting with Ox2h12, further host-range mutants could be isolated on strains resistant to the respective phage: Ox2h12----h12h1----h12h1.1----h12h1.11----h12 h1.111. It was found that Ox2h12h1.1 (and a derivative of Ox2h10, h10h4) probably uses, instead of OmpA or OmpC, yet another outer membrane protein, designated OmpX. Ox2h12h1.11 was obtained on a strain lacking OmpA, -C and -X. This phage could not grow on a mutant of E. coli B, possessing a lipopolysaccharide (LPS) with a defective core oligosaccharide; Ox2h12h1.111 was obtained from this strain. It turned out that the latter two mutants used LPS as a receptor, most likely via its glucose residues. Selection for resistance to them in E. coli B (ompA+, ompC-, ompX-) yielded exclusively LPS mutants, and in another strain, possessing OmpA, C and X, the majority of resistant mutants were of this type. Isolated LPS inactivated the mutant phages very well and was inactive towards Ox2h12. By recombining the genes of mutant phages into the genome of parental phages it could be shown that the phenotypes were associated with gene 38. All mutant alterations (mostly single amino acid substitutions) were found within the hypervariable regions of protein 38. In particular, a substitution leading to Ox2h12h1.11 (Arg170----Ser) had occurred at the same site that led to Ox2h10 (His170----Arg), which binds to OmpC in addition to OmpA. It is concluded that not only can protein 38 gain the ability to switch from a protein to a carbohydrate as a receptor but can do so using the same domain of the polypeptide.
T偶数型大肠杆菌噬菌体Ox2将外膜蛋白OmpA识别为受体。这种识别是由位于病毒体长尾纤维自由末端的266个氨基酸残基的蛋白38完成的。宿主范围突变体是通过三个连续步骤分离得到的：Ox2→Ox2h5→Ox2h10→Ox2h12，其中Ox2h12能有效识别外膜蛋白OmpC，且对OmpA的亲和力有所丧失。与其他噬菌体的这些蛋白相比，蛋白38由两个恒定区和一个连续的四个高变区阵列组成；导致Ox2h12的改变都发生在后者区域内。从Ox2h12开始，可以在对相应噬菌体有抗性的菌株上分离出更多宿主范围突变体：Ox2h12→h12h1→h12h1.1→h12h1.11→h12h1.111。研究发现，Ox2h12h1.1（以及Ox2h10的一个衍生物h10h4）可能不是利用OmpA或OmpC，而是利用另一种外膜蛋白，命名为OmpX。Ox2h12h1.11是在一个缺乏OmpA、OmpC和OmpX的菌株上获得的。这种噬菌体不能在具有缺陷核心寡糖的脂多糖（LPS）的大肠杆菌B突变体上生长；Ox2h12h1.111是从该菌株获得的。结果表明，后两个突变体以LPS为受体，很可能是通过其葡萄糖残基。在大肠杆菌B（ompA +、ompC -、ompX -）中筛选对它们的抗性，只产生了LPS突变体，而在另一个具有OmpA、OmpC和OmpX的菌株中，大多数抗性突变体都是这种类型。分离出的LPS能很好地使突变噬菌体失活，而对Ox2h12没有活性。通过将突变噬菌体的基因重组到亲本噬菌体的基因组中，可以证明这些表型与基因38相关。所有突变改变（大多是单个氨基酸替换）都发生在蛋白38的高变区内。特别是，导致Ox2h12h1.11（Arg170→Ser）的替换发生在与导致Ox2h10（His170→Arg）相同的位点，Ox2h10除了能结合OmpA外还能结合OmpC。得出的结论是，蛋白38不仅能够获得从蛋白质受体转换为碳水化合物受体的能力，而且可以利用多肽的同一结构域来实现这一点。
