Glykos Nicholas M, Papanikolau Yannis, Vlassi Metaxia, Kotsifaki Dina, Cesareni Giovanni, Kokkinidis Michael
Department of Molecular Biology and Genetics, Democritus University of Thrace, Dimitras 19, 68100 Alexandroupolis, Greece.
Biochemistry. 2006 Sep 12;45(36):10905-19. doi: 10.1021/bi060833n.
The repressor of primer (Rop) protein has become a steady source of surprises concerning the relationship between the sequences and the structures of several of its mutants and variants. Here we add another piece to the puzzle of Rop by showing that an engineered deletion mutant of the protein (corresponding to a deletion of residues 30-34 of the wild-type protein and designed to restore the heptad periodicity at the turn region) results in a complete reorganization of the bundle which is converted from a homodimer to a homotetramer. In contrast (and as previously shown), a two-residue insertion, which also restores the heptad periodicity, is essentially identical with wild-type Rop. The new deletion mutant structure is a canonical, left-handed, all-antiparallel bundle with a completely different hydrophobic core and distinct surface properties. The structure agrees and qualitatively explains the results from functional, thermodynamic, and kinetic studies which indicated that this deletion mutant is a biologically inactive hyperstable homotetramer. Additional insight into the stability and dynamics of the mutant structure has been obtained from extensive molecular dynamics simulations in explicit water and with full treatment of electrostatics.
引物阻遏蛋白（Rop）在其多个突变体和变体的序列与结构之间的关系方面，一直不断带来惊喜。在此，我们为Rop之谜增添了新内容，表明该蛋白的一个工程缺失突变体（对应于野生型蛋白30 - 34位残基的缺失，旨在恢复转角区域的七肽周期性）导致了束状结构的完全重组，从同二聚体转变为同四聚体。相比之下（如之前所示），同样恢复七肽周期性的双残基插入突变体与野生型Rop基本相同。新的缺失突变体结构是一种典型的左手全反平行束状结构，具有完全不同的疏水核心和独特的表面性质。该结构与功能、热力学和动力学研究结果相符，并定性地解释了这些结果，这些研究表明该缺失突变体是一种无生物学活性的超稳定同四聚体。通过在明确的水环境中进行广泛的分子动力学模拟并全面处理静电作用，我们对突变体结构的稳定性和动力学有了进一步的了解。
