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首页> 外文期刊>Acta crystallographica, Section F. Structural biology and crystallization communications >Overexpression, crystallization and preliminary X-ray crystallographic analysis of the variable lymphocyte receptor 2913 ectodomain fused with internalin B
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Overexpression, crystallization and preliminary X-ray crystallographic analysis of the variable lymphocyte receptor 2913 ectodomain fused with internalin B

机译：与internalin B融合的可变淋巴细胞受体2913胞外域的过表达，结晶和初步X射线晶体学分析

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摘要

In jawless vertebrates, variable lymphocyte receptors (VLRs) play a crucial role in the recognition of antigens as part of the adaptive immune system. Leucine-rich repeat (LRR) modules and the highly variable insert (HVI) of VLRs contribute to the specificity and diversity of antigen recognition. VLR2913, the antigen of which is not known, contains the same HVI amino-acid sequence as that of VLR RBC36, which recognizes the H-trisaccharide from human blood type O erythrocytes. Since the HVI sequence is rarely identical among all known VLRs, identification of the antigen for VLR2913 and the main contributing factors for antigen recognition based on a comparison of VLR2913 and VLR RBC36 has been attempted. To initiate and facilitate this structural approach, the ectodomain of VLR2913 was fused with the N-terminal domain of internalin B (InlB-VLR2913-ECD). Three amino-acid residues on the concave surface of the LRR modules of InlB-VLR2913-ECD were mutated, considering important residues for hydrogen bonds in the recognition of H-trisaccharide by VLR RBC36. InlB-VLR2913-ECD was overexpressed in Escherichia coli and was crystallized at 295 K using the sitting-drop vapour-diffusion method. X-ray diffraction data were collected to 2.04 angstrom resolution and could be indexed in the tetragonal space group P4(1)2(1)2 (or P4(3)2(1)2), with unit-cell parameters a = 91.12, b = 91.12, c = 62.87 angstrom. Assuming that one monomer molecule was present in the crystallographic asymmetric unit, the calculated Matthews coefficient (V-M) was 2.75 angstrom(3) Da(-1) and the solvent content was 55.2%. Structural determination of InlB-VLR2913-ECD by molecular replacement is in progress.
机译：在无颚脊椎动物中，可变淋巴细胞受体（VLR）在识别抗原中起着至关重要的作用，作为自适应免疫系统的一部分。富含亮氨酸的重复（LRR）模块和VLR的高度可变插入片段（HVI）有助于抗原识别的特异性和多样性。未知抗原的VLR2913含有与VLR RBC36相同的HVI氨基酸序列，该序列识别人血型O红细胞中的H-三糖。由于在所有已知的VLR中HVI序列很少相同，因此已尝试基于VLR2913和VLR RBC36的比较来鉴定VLR2913的抗原和抗原识别的主要因素。为了启动并促进这种结构方法，将VLR2913的胞外域与Internalin B的N端域融合（InlB-VLR2913-ECD）。考虑到在VLR RBC36识别H-三糖中氢键的重要残基，InlB-VLR2913-ECD的LRR模块凹面的三个氨基酸残基发生了突变。 InlB-VLR2913-ECD在大肠杆菌中过表达，并使用坐滴蒸气扩散法在295 K结晶。 X射线衍射数据的采集分辨率为2.04埃，可以在单位空间参数a = 91.12的四边形空间群P4（1）2（1）2（或P4（3）2（1）2）中建立索引。 ，b ＝ 91.12，c ＝ 62.87埃。假设晶体不对称单元中存在一个单体分子，则计算出的马修斯系数（V-M）为2.75埃（3）Da（-1），溶剂含量为55.2％。 InlB-VLR2913-ECD的分子取代结构测定正在进行中。

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