Department of Chemistry, College of the Holy Cross, 1 College Street, Worcester, MA, USA.
Division of Genetics, Wadsworth Center, New York State Department of Health, Albany, NY, 12208, USA.
Sci Rep. 2021 Jun 3;11(1):11680. doi: 10.1038/s41598-021-91090-w.
Protein splicing is a post-translational process by which an intein catalyzes its own excision from flanking polypeptides, or exteins, concomitant with extein ligation. Many inteins have nested homing endonuclease domains that facilitate their propagation into intein-less alleles, whereas other inteins lack the homing endonuclease (HEN) and are called mini-inteins. The mini-intein that interrupts the DNA PolII of Pyrococcus horikoshii has a linker region in place of the HEN domain that is shorter than the linker in a closely related intein from Pyrococcus abyssi. The P. horikoshii PolII intein requires a higher temperature for catalytic activity and is more stable to digestion by the thermostable protease thermolysin, suggesting that it is more rigid than the P. abyssi intein. We solved a crystal structure of the intein precursor that revealed a domain-swapped dimer. Inteins found as domain swapped dimers have been shown to promote intein-mediated protein alternative splicing, but the solved P. horikoshii PolII intein structure has an active site unlikely to be catalytically competent.
蛋白质剪接是一种翻译后过程，其中内含子催化其自身从侧翼多肽（即外显子）中切除，同时外显子连接。许多内含子具有嵌套的归巢内切酶结构域，有助于它们在没有内含子的等位基因中传播，而其他内含子缺乏归巢内切酶（HEN），被称为小型内含子。中断 Pyrococcus horikoshii 的 DNA PolII 的小型内含子在 HEN 结构域的位置上有一个连接区，该连接区比来自 Pyrococcus abyssi 的密切相关内含子的连接区短。P. horikoshii PolII 内含子需要更高的温度才能发挥催化活性，并且对热稳定蛋白酶 thermolysin 的消化更稳定，这表明它比 P. abyssi 内含子更坚固。我们解决了内含子前体的晶体结构，揭示了一个结构域交换的二聚体。已证明作为结构域交换二聚体的内含子可促进内含子介导的蛋白质选择性剪接，但解决的 P. horikoshii PolII 内含子结构具有不太可能具有催化活性的活性位点。
