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X‐ray crystallographic studies of RNase A variants engineered at the most destabilizing positions of the main hydrophobic core: Further insight into protein stability
Author(s) -
Kurpiewska Katarzyna,
Font Josep,
Ribó Marc,
Vilanova Maria,
Lewiński Krzysztof
Publication year - 2009
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/prot.22480
Subject(s) - bovine pancreatic ribonuclease , rnase p , ribonuclease , crystallography , chemistry , protein folding , s tag , protein structure , molecule , folding (dsp implementation) , rnase h , crystal structure , biochemistry , rna , organic chemistry , electrical engineering , gene , engineering
To investigate the structural origin of decreased pressure and temperature stability, the crystal structure of bovine pancreatic ribonuclease A variants V47A, V54A, V57A, I81A, I106A, and V108A was solved at 1.4–2.0 Å resolution and compared with the structure of wild‐type protein. The introduced mutations had only minor influence on the global structure of ribonuclease A. The structural changes had individual character that depends on the localization of mutated residue, however, they seemed to expand from mutation site to the rest of the structure. Several different parameters have been evaluated to find correlation with decrease of free energy of unfolding ΔΔG T , and the most significant correlation was found for main cavity volume change. Analysis of the difference distance matrices revealed that the ribonuclease A molecule is organized into five relatively rigid subdomains with individual response to mutation. This behavior consistent with results of unfolding experiments is an intrinsic feature of ribonuclease A that might be surviving remnants of folding intermediates and reflects the dynamic nature of the molecule. Proteins 2009. © 2009 Wiley‐Liss, Inc.