Bioinformatic method for protein thermal stabilization by structural entropy optimization

Abstract

Engineering proteins for higher thermal stability is an important and difficult challenge. We describe a bioinformatic method incorporating sequence alignments to redesign proteins to be more stable through optimization of local structural entropy. Using this method, improved configurational entropy (ICE), we were able to design more stable variants of a mesophilic adenylate kinase with only the sequence information of one psychrophilic homologue. The redesigned proteins display considerable increases in their thermal stabilities while still retaining catalytic activity. ICE does not require a three-dimensional structure or a large number of homologous sequences, indicating a broad applicability of this method. Our results also highlight the importance of entropy in the stability of protein structures.adenylate kinase ͉ improved configurational entropy ͉ local structural entropy ͉ protein engineering ͉ protein stability

Crystal structures of thermally stable adenylate kinase mutants designed by local structural entropy optimization and structure-guided mutagenesis

“…LSE is an empirical descriptor of conformational variability in a protein sequence (Chan et al, 2004) that is computed based on structural information derived from the Protein Data Bank (PDB) (Berman et al, 2000). Reducing the LSE of a protein sequence by introducing mutations can result in fewer conformational states and thus a more stable structure (Bae et al, 2008). This approach does not require structural information of the target proteins but is limited by the fact that LSE overlooks and can damage global stabilizing structural features connecting distant regions of a polypeptide (Moon et al, 2014a).…”

“…We previously developed a stabilization method based on measures of local structural entropy (LSE) (Bae et al, 2008). LSE is an empirical descriptor of conformational variability in a protein sequence (Chan et al, 2004) that is computed based on structural information derived from the Protein Data Bank (PDB) (Berman et al, 2000).…”

Crystal structure of a trimeric archaeal adenylate kinase from the mesophile Methanococcus maripaludis with an unusually broad functional range and thermal stability

“…Even a transient loss of AK function leads to a loss of cell viability and death 1. In addition to its physiological importance, AK has also served as a model for studies in molecular evolution,2, 3 protein folding, and dynamics 4–13. There are two oligomeric classes of AK: monomeric as found in Eubacteria and trimeric as found in Archaebacteria.…”

Improving the Thermostability and Activity of a Thermophilic Subtilase by Incorporating Structural Elements of Its Psychrophilic Counterpart

“…However, there is evidence showing that structural components that are involved in the catalytic cycle have improved flexibility to increase low-temperature activity of psychrophilic enzymes, while other protein regions that are not implicated in catalysis may be even more rigid than their mesophilic counterparts (13-15). Indeed, thermostable variants of thermophilic and mesophilic proteins have been successfully constructed by incorporating structural elements of their less stable counterparts (16,17).…”

“…However, there is evidence showing that structural components that are involved in the catalytic cycle have improved flexibility to increase low-temperature activity of psychrophilic enzymes, while other protein regions that are not implicated in catalysis may be even more rigid than their mesophilic counterparts (13-15). Indeed, thermostable variants of thermophilic and mesophilic proteins have been successfully constructed by incorporating structural elements of their less stable counterparts (16,17).Subtilisin-like serine proteases (subtilases) have been extensively studied due to their contributions to the understanding of the mechanism of enzyme catalysis as well as their industrial potential (18). Thermostability is one of the main requirements for commercial enzymes, and enzymes with high catalytic activity at low temperatures offer economic and environmental benefits through energy savings (6,19,20).…”