2LCJ

Solution NMR structure of Pab PolII Intein

2LCJ の概要

• エントリーDOI: 10.2210/pdb2lcj/pdb
• NMR情報: BMRB: 17418
• 分子名称: Pab polC intein (1 entity in total)
• 機能のキーワード: hydrolase
• 由来する生物種: Pyrococcus abyssi
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 21553.29

構造登録者

Jiajing, L.,Mills, K.V.,Albracht, C.D. (登録日: 2011-04-29, 公開日: 2011-09-21, 最終更新日: 2024-05-15)

主引用文献

Du, Z.,Liu, J.,Albracht, C.D.,Hsu, A.,Chen, W.,Marieni, M.D.,Colelli, K.M.,Williams, J.E.,Reitter, J.N.,Mills, K.V.,Wang, C.
Structural and Mutational Studies of a Hyperthermophilic Intein from DNA Polymerase II of Pyrococcus abyssi.
J.Biol.Chem., 286:38638-38648, 2011

PubMed Abstract: Protein splicing is a precise self-catalyzed process in which an intein excises itself from a precursor with the concomitant ligation of the flanking polypeptides (exteins). Protein splicing proceeds through a four-step reaction but the catalytic mechanism is not fully understood at the atomic level. We report the solution NMR structures of the hyperthermophilic Pyrococcus abyssi PolII intein, which has a noncanonical C-terminal glutamine instead of an asparagine. The NMR structures were determined to a backbone root mean square deviation of 0.46 Å and a heavy atom root mean square deviation of 0.93 Å. The Pab PolII intein has a common HINT (hedgehog intein) fold but contains an extra β-hairpin that is unique in the structures of thermophilic inteins. The NMR structures also show that the Pab PolII intein has a long and disordered loop in place of an endonuclease domain. The N-terminal Cys-1 amide is hydrogen bonded to the Thr-90 hydroxyl in the conserved block-B TXXH motif and the Cys-1 thiol forms a hydrogen bond with the block F Ser-166. Mutating Thr-90 to Ala dramatically slows N-terminal cleavage, supporting its pivotal role in promoting the N-S acyl shift. Mutagenesis also showed that Thr-90 and His-93 are synergistic in catalyzing the N-S acyl shift. The block F Ser-166 plays an important role in coordinating the steps of protein splicing. NMR spin relaxation indicates that the Pab PolII intein is significantly more rigid than mesophilic inteins, which may contribute to the higher optimal temperature for protein splicing.

PubMed: 21914805
DOI: 10.1074/jbc.M111.290569

実験手法

SOLUTION NMR