3AFG

Crystal structure of ProN-Tk-SP from Thermococcus kodakaraensis

3AFG の概要

• エントリーDOI: 10.2210/pdb3afg/pdb
• 関連するPDBエントリー: 1SCJ
• 分子名称: Subtilisin-like serine protease, CALCIUM ION (3 entities in total)
• 機能のキーワード: subtilisin, propeptide, thermococcus kodakaraensis, hydrolase, protease, serine protease
• 由来する生物種: Thermococcus kodakarensis
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 113970.69

構造登録者

Foophow, T.,Tanaka, S.,Angkawidjaja, C.,Koga, Y.,Takano, K.,Kanaya, S. (登録日: 2010-03-01, 公開日: 2010-08-04, 最終更新日: 2023-11-01)

主引用文献

Foophow, T.,Tanaka, S.,Angkawidjaja, C.,Koga, Y.,Takano, K.,Kanaya, S.
Crystal structure of a subtilisin homologue, Tk-SP, from Thermococcus kodakaraensis: requirement of a C-terminal beta-jelly roll domain for hyperstability.
J.Mol.Biol., 400:865-877, 2010

PubMed Abstract: Tk-SP is a hyperthermostable subtilisin-like serine protease from Thermococcus kodakaraensis and is autoprocessed from its precursor (Pro-Tk-SP) with N- and C-propeptides. The crystal structure of the active-site mutant of Pro-Tk-SP lacking C-propeptide, ProN-Tk-S359A, was determined at 2.0 A resolution. ProN-Tk-S359A consists of the N-propeptide, subtilisin, and beta-jelly roll domains. Two Ca(2+) ions bind to the beta-jelly roll domain. The overall structure of ProN-Tk-S359A without the beta-jelly roll domain is similar to that of the bacterial propeptide:subtilisin complex, except that it does not contain Ca(2+) ions. To analyze the role of the beta-jelly roll domain of Tk-SP, we constructed a series of the active-site mutants of Tk-SP with (Tk-S359A/C) and without (Tk-S359A/CDeltaJ) beta-jelly roll domain. Both Tk-S359C and Tk-S359CDeltaJ exhibited protease activities in gel assay, indicating that the beta-jelly roll domain is not required for folding or activity. However, the T(m) value of Tk-S359ADeltaJ determined by far-UV CD spectroscopy in the presence of 10-mM CaCl(2) was lower than that of Tk-S359A by 29.4 degrees C. The T(m) value of Tk-S359A was decreased by 29.5 degrees C by the treatment with 10 mM ethylenediaminetetraacetic acid, indicating that the beta-jelly roll domain contributes to the stabilization of Tk-S359A only in a Ca(2+)-bound form. Tk-SP highly resembles subtilisin-like serine proteases from Pyrococcus furiosus, Thermococcus gammatolerans, and Thermococcus onnurineus in size and amino acid sequence. We propose that attachment of a beta-jelly roll domain to the C-terminus is one of the strategies of the proteins from hyperthermophiles to adapt to high-temperature environment.

PubMed: 20595040
DOI: 10.1016/j.jmb.2010.05.064

実験手法

X-RAY DIFFRACTION (2 Å)