1HZC

BACILLUS CALDOLYTICUS COLD-SHOCK PROTEIN MUTANTS TO STUDY DETERMINANTS OF PROTEIN STABILITY

1HZC の概要

• エントリーDOI: 10.2210/pdb1hzc/pdb
• 関連するPDBエントリー: 1C9O 1CSP 1HZ9 1HZA 1HZB
• 分子名称: COLD SHOCK PROTEIN CSPB, SODIUM ION (3 entities in total)
• 機能のキーワード: beta barrel, homodimer, transcription
• 由来する生物種: Bacillus caldolyticus
• タンパク質・核酸の鎖数: 2
• 化学式量合計: 14565.04

構造登録者

Delbrueck, H.,Mueller, U.,Perl, D.,Schmid, F.X.,Heinemann, U. (登録日: 2001-01-24, 公開日: 2001-11-07, 最終更新日: 2023-08-09)

主引用文献

Delbruck, H.,Mueller, U.,Perl, D.,Schmid, F.X.,Heinemann, U.
Crystal structures of mutant forms of the Bacillus caldolyticus cold shock protein differing in thermal stability.
J.Mol.Biol., 313:359-369, 2001

PubMed Abstract: The cold shock proteins Bc-Csp from the thermophile Bacillus caldolyticus and Bs-CspB from the mesophile Bacillus subtilis differ significantly in their conformational stability, although the two proteins differ by only 12 out of 67 amino acid residues. The three-dimensional structure of these small and compact beta-barrel proteins without disulfide bonds, cis-proline residues or tightly bound cofactors is very similar. Previous work has shown that Bc-Csp displays a twofold increase in the free energy of stabilization relative to its homolog Bs-CspB, and indicated that electrostatic interactions are, in part, responsible for this effect. It was further described that the stability difference is almost exclusively due to surface-exposed charged residues at sequence positions 3 and 66 of Bc-Csp and Bs-CspB, whereas all other amino acid changes between both proteins have no net effect on stability. To investigate how two surface residues determine the stability of Bc-Csp, Arg3 and Leu66 were replaced by glutamic acid, corresponding to the Bs-CspB sequence. The crystal structures of the resultant protein variants, Bc-Csp R3E and Bc-Csp L66E, were determined at 1.4 A and 1.27 A resolution, and refined to R values of 13.9 % and 15.8 %, respectively. Both structures closely resemble Bc-Csp in their global fold and show different hydrogen bonding and salt-bridge patterns when two independent molecules in the asymmetric unit of the crystal are compared. To extend the study to neighbored residues that help determine the surface charge around Arg3 and Leu66, the mutant proteins Bc-Csp E46A, Bc-Csp R3E/E46A/L66E and Bc-Csp V64T/L66E/67A were crystallized. Their structures were determined at resolutions of 1.8 A, 1.32 A and 1.8 A and refined to R values of 18.5 %, 13.8 % and 19.3 %, respectively. A systematic comparison of the crystal structures of all forms of the B. caldolyticus cold shock protein shows varying patterns of hydrogen bonds and electrostatic interactions around residues 3 and 66. Thermal destabilization of the protein by mutation appears to correlate with the extent of an acidic surface patch near the C-terminal carboxylate group.

PubMed: 11800562
DOI: 10.1006/jmbi.2001.5051

実験手法

X-RAY DIFFRACTION (1.32 Å)