Nordlund Henri R, Hytönen Vesa P, Hörhä Jarno, Määttä Juha A E, White Daniel J, Halling Katrin, Porkka Eevaleena J, Slotte J Peter, Laitinen Olli H, Kulomaa Markku S
Department of Biological and Environmental Science, NanoScience Center, P.O. Box 35 (YAB), FIN-40014 University of Jyväskylä, Finland.
Biochem J. 2005 Dec 15;392(Pt 3):485-91. doi: 10.1042/BJ20051038.
scAvd (single-chain avidin, where two dcAvd are joined in a single polypeptide chain), having four biotin-binding domains, was constructed by fusion of topologically modified avidin units. scAvd showed similar biotin binding and thermal stability properties as chicken avidin. The DNA construct encoding scAvd contains four circularly permuted avidin domains, plus short linkers connecting the four domains into a single polypeptide chain. In contrast with wild-type avidin, which contains four identical avidin monomers, scAvd enables each one of the four avidin domains to be independently modified by protein engineering. Therefore the scAvd scaffold can be used to construct spatially and stoichiometrically defined pseudotetrameric avidin molecules showing different domain characteristics. In addition, unmodified scAvd could be used as a fusion partner, since it provides a unique non-oligomeric structure, which is fully functional with four high-affinity biotin-binding sites. Furthermore, the subunit-to-domain strategy described in the present study could be applied to other proteins and protein complexes, facilitating the development of sophisticated protein tools for applications in nanotechnology and life sciences.
单链抗生物素蛋白（scAvd，由两个双链抗生物素蛋白连接在一条多肽链中组成）有四个生物素结合结构域，它通过拓扑修饰的抗生物素蛋白单元融合构建而成。scAvd表现出与鸡抗生物素蛋白相似的生物素结合和热稳定性特性。编码scAvd的DNA构建体包含四个环状排列的抗生物素蛋白结构域，以及将这四个结构域连接成一条多肽链的短连接子。与含有四个相同抗生物素蛋白单体的野生型抗生物素蛋白不同，scAvd能使四个抗生物素蛋白结构域中的每一个都通过蛋白质工程进行独立修饰。因此，scAvd支架可用于构建具有不同结构域特征的空间和化学计量定义的假四聚体抗生物素蛋白分子。此外，未修饰的scAvd可作为融合伙伴，因为它提供了一种独特的非寡聚结构，该结构具有四个高亲和力生物素结合位点且功能完整。此外，本研究中描述的亚基到结构域策略可应用于其他蛋白质和蛋白质复合物，有助于开发用于纳米技术和生命科学应用的精密蛋白质工具。
