Pardo-Avila Fátima, Kudva Renuka, Levitt Michael, von Heijne Gunnar
Department of Structural Biology, Stanford University, Palo Alto, CA, USA.
Department of Biochemistry and Biophysics, Stockholm University, SE-106 91 Stockholm, Sweden.
bioRxiv. 2024 Aug 20:2024.08.20.608737. doi: 10.1101/2024.08.20.608737.
Nascent polypeptide chains (NCs) are extruded from the ribosome through an exit tunnel (ET) traversing the large ribosomal subunit. The ET's irregular and chemically complex wall allows for various NC-ET interactions. Translational arrest peptides (APs) bind in the ET to induce translational arrest, a property that can be exploited to study NC-ET interactions by Force Profile Analysis (FPA). We employed FPA and molecular dynamics (MD) simulations to investigate how individual residues placed in a glycine-serine repeat segment within an AP-stalled NC interact with the ET to exert a pulling force on the AP and release stalling. Our results indicate that large and hydrophobic residues generate a pulling force on the NC when placed ≳10 residues away from the peptidyl transfer center (PTC). Moreover, an asparagine placed 12 residues from the PTC makes a specific stabilizing interaction with the tip of ribosomal protein uL22 that reduces the pulling force on the NC, while a lysine or leucine residue in the same position increases the pulling force. Finally, the MD simulations suggest how the SecM AP interacts with the ET to promote translational stalling.
新生多肽链（NCs）通过贯穿核糖体大亚基的出口通道（ET）从核糖体中挤出。ET的不规则且化学组成复杂的壁允许各种NC-ET相互作用。翻译抑制肽（APs）结合在ET中以诱导翻译抑制，这一特性可用于通过力谱分析（FPA）研究NC-ET相互作用。我们采用FPA和分子动力学（MD）模拟来研究置于AP停滞的NC内甘氨酸-丝氨酸重复片段中的单个残基如何与ET相互作用，从而对AP施加拉力并解除停滞。我们的结果表明，当置于距肽基转移中心（PTC）≳10个残基处时，大的疏水残基会对NC产生拉力。此外，距PTC 12个残基处的天冬酰胺与核糖体蛋白uL22的末端形成特定的稳定相互作用，从而降低对NC的拉力，而相同位置的赖氨酸或亮氨酸残基则会增加拉力。最后，MD模拟表明SecM AP如何与ET相互作用以促进翻译停滞。
