Thamotharan Subbiah, Karthikeyan Tangavelou, Kulkarni Kiran A, Shetty Kartika N, Surolia Avadhesha, Vijayan Mamannamana, Suguna Kaza
Molecular Biophysics Unit, Indian Institute of Science, Bangalore, India.
Acta Crystallogr D Biol Crystallogr. 2011 Mar;67(Pt 3):218-27. doi: 10.1107/S0907444911004525. Epub 2011 Feb 15.
A mutant of Erythrina corallodendron lectin was generated with the aim of enhancing its affinity for N-acetylgalactosamine. A tyrosine residue close to the binding site of the lectin was mutated to a glycine in order to facilitate stronger interactions between the acetamido group of the sugar and the lectin which were prevented by the side chain of the tyrosine in the wild-type lectin. The crystal structures of this Y106G mutant lectin in complex with galactose and N-acetylgalactosamine have been determined. A structural rationale has been provided for the differences in the relative binding affinities of the wild-type and mutant lectins towards the two sugars based on the structures. A hydrogen bond between the O6 atom of the sugars and the variable loop of the carbohydrate-binding site of the lectin is lost in the mutant complexes owing to a conformational change in the loop. This loss is compensated by an additional hydrogen bond that is formed between the acetamido group of the sugar and the mutant lectin in the complex with N-acetylgalactosamine, resulting in a higher affinity of the mutant lectin for N-acetylgalactosamine compared with that for galactose, in contrast to the almost equal affinity of the wild-type lectin for the two sugars. The structure of a complex of the mutant with a citrate ion bound at the carbohydrate-binding site that was obtained while attempting to crystallize the complexes with sugars is also presented.
为了增强刺桐凝集素对N-乙酰半乳糖胺的亲和力，构建了一种刺桐凝集素突变体。将靠近凝集素结合位点的一个酪氨酸残基突变为甘氨酸，以促进糖的乙酰氨基基团与凝集素之间形成更强的相互作用，而野生型凝集素中的酪氨酸侧链阻碍了这种相互作用。已确定该Y106G突变体凝集素与半乳糖和N-乙酰半乳糖胺复合物的晶体结构。基于这些结构，为野生型和突变型凝集素对两种糖的相对结合亲和力差异提供了结构上的解释。由于环的构象变化，在突变体复合物中糖的O6原子与凝集素碳水化合物结合位点的可变环之间的氢键消失。这种损失由糖的乙酰氨基基团与N-乙酰半乳糖胺复合物中的突变型凝集素之间形成的额外氢键补偿，导致突变型凝集素对N-乙酰半乳糖胺的亲和力高于对半乳糖的亲和力，这与野生型凝集素对两种糖几乎相等的亲和力形成对比。还展示了在尝试用糖结晶复合物时获得的突变体与结合在碳水化合物结合位点的柠檬酸根离子的复合物结构。
