6FRG

Crystal structure of G-1F mutant of Ssp DnaB Mini-Intein variant M86

6FRG の概要

• エントリーDOI: 10.2210/pdb6frg/pdb
• 分子名称: Replicative DNA helicase, PENTAETHYLENE GLYCOL, TRIETHYLENE GLYCOL, ... (5 entities in total)
• 機能のキーワード: intein, protein splicing, hint domain fold, pre-splicing form, splicing
• 由来する生物種: Synechocystis sp. (strain PCC 6803 / Kazusa); 詳細
• タンパク質・核酸の鎖数: 1
• 化学式量合計: 20028.09

構造登録者

Popp, M.A.,Blankenfeldt, W.,Friedel, K.,Mootz, H.D. (登録日: 2018-02-15, 公開日: 2019-01-30, 最終更新日: 2024-01-17)

主引用文献

Friedel, K.,Popp, M.A.,Matern, J.C.J.,Gazdag, E.M.,Thiel, I.V.,Volkmann, G.,Blankenfeldt, W.,Mootz, H.D.
A functional interplay between intein and extein sequences in protein splicing compensates for the essential block B histidine.
Chem Sci, 10:239-251, 2019

PubMed Abstract: Inteins remove themselves from a precursor protein by protein splicing. Due to the concomitant structural changes of the host protein, this self-processing reaction has enabled many applications in protein biotechnology and chemical biology. We show that the evolved M86 mutant of the DnaB intein displays a significantly improved tolerance towards non-native amino acids at the N-terminally flanking (-1) extein position compared to the parent intein, in the form of both an artificially -splicing split intein and the -splicing mini-intein. Surprisingly, side chains with increased steric bulk compared to the native Gly(-1) residue, including d-amino acids, were found to compensate for the essential block B histidine in His73Ala mutants in the initial N-S acyl shift of the protein splicing pathway. In the case of the M86 intein, large (-1) side chains can even rescue protein splicing activity as a whole. With the comparison of three crystal structures, namely of the M86 intein as well as of its Gly(-1)Phe and Gly(-1)Phe/His73Ala mutants, our data supports a model in which the intein's active site can exert a strain by varying mechanisms on the different angles of the scissile bond at the extein-intein junction to effect a ground-state destabilization. The compensatory mechanism of the block B histidine is the first example for the direct functional role of an extein residue in protein splicing. It sheds new light on the extein-intein interplay and on possible consequences of their co-evolution as well as on the laboratory engineering of improved inteins.

PubMed: 30713635
DOI: 10.1039/c8sc01074a

実験手法

X-RAY DIFFRACTION (1.535 Å)