The antibodies directed against N-terminal heptad-repeat peptide of hRSV fusion protein and its analog-5-Helix inhibit virus infection in vitro

Human respiratory syncytial virus (hRSV) membrane fusion is promoted by the formation of a trimer-of-hairpins structure that brings the amino- and carboxyl-terminal regions of fusion (F) protein into close proximity. Two heptad-repeat (HR1 and HR2) regions in F protein play an important role in this process. Our previous study demonstrated that peptides derived from HR1 and HR2 regions of F protein were potent inhibitors of hRSV entry. Here we showed that HR1 peptide and its analog denoted 5-Helix which contained a central coiled-coil formed by three HR1s could induce highly potent antibody response in the immunized rabbits. Both antibodies could recognize F1 domain of the F protein and inhibited hRSV entry with the neutralizing antibody titers of 1:61 and 1:115, respectively. These suggested that 5-Helix could induce potent neutralizing antibody response and the central coiled-coil might be a highly conserved neutralization site for hRSV F protein.     

细胞动粒蛋白Hec1启动子的结构及功能分析

细胞的分裂是一个严格调控,高度有序的过程.为了将复制后的染色体均匀、准确地传递给两个子细胞,细胞在分裂中后期受到纺锤体检验点的严格监控.Hec1定位于动粒,是纺锤体检验点调控的关键蛋白之一,它通过螺旋 螺旋结构域与其他动粒蛋白相互作用调节姐妹染色体的精确分离.为研究Hec1转录水平的调控机理,采用BLAST工具,从GenBank 中搜索到了人Hec1基因上游的序列,并利用在线工具http://mbs.cbrc.jp/research/db/TFSEARCH.html提供的转录因子结合位点搜索引擎,对其5′启动子调节区段进行了分析.分析结果表明：在Hec1基因上游-200～-1序列内,存在E2F、ATF4和cAMP应答元件结合蛋白（CREB）等转录因子调控元件.在结构分析的基础上,提取HeLa细胞基因组DNA,用PCR方法克隆了Hec1基因启动子,并构建了多个含启动子不同区段的pGL3荧光素酶报告基因表达质粒.瞬时转染HeLa细胞后的结果表明,-70～－63以及－155～－144之间的启动子区对维持荧光素酶活性最为关键.凝胶迁移实验证明,这两个区段分别能够和转录因子CREB以及ATF4结合.随后,采用野生型的以及含有133位磷酸化位点突变的CREB转染HeLa细胞,通过荧光定量PCR实验发现,Hec1的表达水平分别出现明显上升和下降.该结果表明,Hec1表达的调控是通过CREB的活化来完成的.     

Crystal structure of a coiled-coil domain from human ROCK I

The small GTPase Rho and one of its targets, Rho-associated kinase (ROCK), participate in a variety of actin-based cellular processes including smooth muscle contraction, cell migration, and stress fiber formation. The ROCK protein consists of an N-terminal kinase domain, a central coiled-coil domain containing a Rho binding site, and a C-terminal pleckstrin homology domain. Here we present the crystal structure of a large section of the central coiled-coil domain of human ROCK I (amino acids 535-700). The structure forms a parallel α-helical coiled-coil dimer that is structurally similar to tropomyosin, an actin filament binding protein. There is an unusual discontinuity in the coiled-coil; three charged residues (E613, R617 and D620) are positioned at what is normally the hydrophobic core of coiled-coil packing. We speculate that this conserved irregularity could function as a hinge that allows ROCK to adopt its autoinhibited conformation.