Aromatic clusters: a determinant of thermal stability of thermophilic proteins

Abstract

A number of factors have been elucidated as responsible for the thermal stability of thermophilic proteins. However, the contribution of aromatic interactions to thermal stability has not been systematically studied. In the present investigation we used a graph spectral method to identify aromatic clusters in a dataset of 24 protein families for which the crystal structures of both the thermophilic and their mesophilic homologues are known. Our analysis shows a presence of additional aromatic clusters or enlarged aromatic networks in 17 different thermophilic protein families, which are absent in the corresponding mesophilic homologue. The additional aromatic clusters identified in the thermophiles are smaller in size and are largely found on the protein surface. The aromatic clusters are found to be relatively rigid regions of the surface and often the additional aromatic cluster is located close to the active site of the thermophilic enzyme. The residues in the additional aromatic clusters are preferably mutated to Leu, Ser or Ile in the mesophilic homologue. An analysis of the packing geometry of the pairwise aromatic interaction in the additional aromatic clusters shows a preference for a T-shaped orthogonal packing geometry. The present study also provides new insights for protein engineers to design thermostable and thermophilic proteins.

Amino acid contacts in proteins adapted to different temperatures: hydrophobic interactions and surface charges play a key role

“…Using a higher threshold than 6Å , would result in inclusion of too many residue pairs and reflect the general environment around the amino acid rather than specific contacts. We are most likely to miss contacts between large amino acids like Arg, Lys and Trp, possibly explaining why we find little tendency for the aromatic clustering reported by Kannan and Vishveshwara (2000). While we may have missed some contacts, we think we have captured the major differences between the proteins in terms of changes in sidechain contacts.…”

“…Kannan and Vishveshwara (2000) suggested that aromatic clusters play a key role in thermostabilization. We cannot see any of these trends in our data, even though Tyr, Arg and Lys are more common in protein from species adapted to the highest temperatures.…”

“…Histidine was also placed in a separate category. We also made a category of Tyr and Trp, since these amino acids are important for forming connective networks, as have been suggested by Kannan andVishveshwara 2000 andBrinda andVishveshwara et al 2005.…”

“…The aromatic amino acids have been theorised to play an important role in thermostabilisation, and many interaction types involving aromatic amino acids have been dicussed. Kannan and Vishveshwara (2000) suggested that aromatic clusters could play an important role in thermostabilisation. If this were the case, one would expect our analysis to show an increase in aromatic-aromatic contacts.…”

“…Cation-p bonds that form between the positively charged residues arginine and lysine and the aromatic residues, particularly Tyr and Trp, have been suggested to be of importance in thermostabilisation by Chakravarty and Varadarajan (2002) as well as Gromiha et al (2002). Kannan and Vishveshwara (2000) suggested that aromatic clusters are important in thermostabilisation.…”