Abstract：
Spectroscopic studies on domains and peptides of large proteins are complicated because of the tendency of short peptides to form oligomers in aquatic buffers, but conjugation of a peptide with a carrier protein may be helpful. In this study we approved that a fragment of SK30 peptide from phospholipase A2 domain of VP1 Parvovirus B19 capsid protein (residues: 144-159; 164; 171-183; sequence: SAVDSAARIHDFRYSQLAKLGINPYTHWTVADEELLKNIK) turns from random coil to alpha helix in the acidic medium only in case if it had been conjugated with BSA (through additional N-terminal Cys residue, turning it into CSK31 peptide, and SMCC linker) according to CD-spectroscopy results. In contrast, unconjugated SK30 peptide does not undergo such shift because it forms stable oligomers connected by intermolecular antiparallel beta sheet, according to IR-spectroscopy, CD-spectroscopy, blue native gel electrophoresis and centrifugal ultrafiltration, as, probably, the whole isolated phospholipase domain of VP1 protein does. However, being a part of the long VP1 capsid protein, phospholipase domain may change its fold during the acidification of the medium in the endolysosome by the way of the formation of contacts between protonated His153 and Asp175, promoting the shift from random coil to alpha helix in its N-terminal part. This study opens up a perspective of vaccine development, since rabbit polyclonal antibodies against the conjugate of CSK31 peptide with BSA, in which the structure of the second alpha helix from the phospholipase A2 domain should be reproduced, can bind epitopes of the complete recombinant unique part of VP1 Parvovirus B19 capsid (residues: 1-227).
摘要：
大蛋白的结构域和肽的光谱研究是复杂的，因为短肽在水生缓冲液中形成寡聚体的趋势，但是肽与载体蛋白的结合可能会有所帮助。在这项研究中，我们批准了来自VP1细小病毒B19衣壳蛋白的磷脂酶A2结构域的SK30肽片段（残基：144-159;164;171-183;序列：SAVDSAARIHDFRYSQLAKLGINPYTHWTVADEELLKNIK）在酸性介质中从无规卷曲变成α螺旋（如果它与BSA末端残基通过额外的N偶联，把它变成CSK31肽，和SMCC接头）根据CD光谱学结果。相比之下,未缀合的SK30肽不经历这种转变，因为它形成由分子间反平行β折叠连接的稳定寡聚体，根据红外光谱，CD-光谱学,蓝色天然凝胶电泳和离心超滤，as,可能,VP1蛋白的整个分离的磷脂酶结构域。然而,作为长VP1衣壳蛋白的一部分，磷脂酶结构域可能会在内溶酶体中培养基酸化过程中，通过质子化的His153和Asp175之间形成接触的方式改变其折叠，从而促进其N末端部分从无规卷曲向α螺旋的转变。这项研究开辟了疫苗开发的前景，由于兔针对CSK31肽与BSA缀合物的多克隆抗体，其中应该复制磷脂酶A2结构域的第二个α螺旋的结构，可以结合VP1细小病毒B19衣壳的完整重组独特部分的表位(残基:1-227)。
