Crystallization of a trimeric human T cell leukemia virus type 1 gp21 ectodomain fragment as a chimera with maltose-binding protein
Crystallization of a trimeric human T cell leukemia virus type 1 gp21 ectodomain fragment as a chimera with maltose-binding protein
复制标题
DOI：
10.1002/pro.5560070715
复制
发表时间：
1998-07-01
期刊：
影响因子：
8
通讯作者：
Poumbourios, P
中科院分区：
文献类型：
--
作者：
Center, RJ;Kobe, B;Poumbourios, P
我们提出了一种新型蛋白质结晶策略，应用于缺乏融合肽和跨膜结构域的人T细胞白血病病毒1型（HTLV-1）跨膜蛋白gp 21的结晶，作为与大肠杆菌麦芽糖结合蛋白（MBP）的嵌合体。晶体不能用MBP/gp 21融合蛋白获得，其中融合伴侣被柔性接头分开，但在通过三个丙氨酸残基将MBP C-末端α-螺旋连接到gp 21的预测N-末端α-螺旋序列后获得。通过沉降平衡和X-射线衍射评估，gp 21序列赋予可溶性融合蛋白三聚体结构，与其他逆转录病毒跨膜蛋白的三聚体结构一致。包膜蛋白前体gp 62在哺乳动物细胞中表达时同样是三聚体。我们的研究结果表明，MBP可能有一个一般的应用程序，含有N-末端α-螺旋序列的蛋白质的结晶。
We present a novel protein crystallization strategy, applied to the crystallization of human T cell leukemia virus type 1 (HTLV-1) transmembrane protein gp21 lacking the fusion peptide and the transmembrane domain, as a chimera with the Escherichia coli maltose binding protein (MBP). Crystals could not be obtained with a MBP/gp21 fusion protein in which fusion partners were separated by a flexible linker, but were obtained after connecting the MBP C-terminal alpha-helix to the predicted N-terminal alpha-helical sequence of gp21 via three alanine residues. The gp21 sequences conferred a trimeric structure to the soluble fusion proteins as assessed by sedimentation equilibrium and X-ray diffraction, consistent with the trimeric structures of other retroviral transmembrane proteins. The envelope protein precursor, gp62, is likewise trimeric when expressed in mammalian cells. Our results suggest that MBP may have a general application for the crystallization of proteins containing N-terminal alpha-helical sequences.