3a is an accessory protein from SARS coronavirus that is known to play a significant role in the proliferation of the virus by forming tetrameric ion channels. Although the monomeric units are known to consist of three transmembrane (TM) domains, there are no solved structures available for the complete monomer. The present study proposes a structural model for the transmembrane region of the monomer by employing our previously tested approach, which predicts potential orientations of TM 𝛼-helices by minimizing the unfavorable contact surfaces between the different TM domains. The best model structure comprising all three 𝛼-helices has been subjected to MD simulations to examine its quality. The TM bundle was found to form a compact and stable structure with significant intermolecular interactions. The structural features of the proposed model of 3a account for observations from previous experimental investigations on the activity of the protein. Further analysis indicates that residues from the TM2 and TM3 domains are likely to line the pore of the ion channel, which is in good agreement with a recent experimental study. In the absence of an experimental structure for the protein, the proposed structure can serve as a useful model for inferring structure-function relationships about the protein.

An understanding of the determinants of the thermal stability of thermostable proteins is expected to enable design of enzymes that can be employed in industrial biocatalytic processes carried out at high temperatures. A major factor that has been proposed to stabilize thermostable proteins is the high occurrenceof salt bridges. The current study employs free energy calculations to elucidate the thermodynamics of the formation of salt bridge interactions and the temperature dependence, using acetate and methylguanidium ionsas model systems. Three different orientations of the methylguanidinium approaching the carboxylate grouphave been considered for obtaining the free energy profiles. The association of the two ions becomes more favorable with an increase in temperature. The desolvation penalty corresponding to the association of the ionpair is the lowest at high temperatures. The occurrence of bridging water molecules between the ions ensures that the ions are not fully desolvated, and this could provide an explanation for the existence of internal watermolecules in thermostable proteins reported recently. The findings provide a detailed picture of the interactions that make ion pair association at high temperatures a favorable process, and reaffirm the importance of saltbridges in the design of thermostable proteins.