Intrinsically disordered regions (IDRs) and protein (IDPs) are highly flexible owing to their lack of well-defined structures. A subset of such proteins interacts with various substrates; including RNA; frequently adopting regular structures in the final complex. In this work; we have analysed a dataset of protein–RNA complexes undergoing disorder-to-order transition (DOT) upon binding. We found that DOT regions are generally small in size (less than 3 residues) for RNA binding proteins. Like structured proteins; positively charged residues are found to interact with RNA molecules; indicating the dominance of electrostatic and cation-π interactions. However, a comparison of binding frequency shows that interface hydrophobic and aromatic residues have more interactions in only DOT regions than in a protein. Further; DOT regions have significantly higher exposure to water than their structured counterparts. Interactions of DOT regions with RNA increase the sheet formation with minor changes in helix forming residues. We have computed the interaction energy for amino acids–nucleotide pairs; which showed the preference of His–G; Asn–U and Ser–U at for the interface of DOT regions. This study provides insights to understand protein–RNA interactions and the results could also be used for developing a tool for identifying DOT regions in RNA binding proteins.

Protein—RNA complexes play important roles in various biological processes. The functions of protein—RNA complexes are dictated by their interactions, binding, stability and affinity. In this work, we have identified the key residues (KRs), which are involved in both stability and binding. We found that 42% of considered proteins share common binding and stabilizing residues, whereas these residues are distinct in 58% of the proteins. Overall, 5% of stabilizing and 3% of binding residues serve as key residues. These residues are enriched with the combination of polar, charged, aliphatic and aromatic residues. In addition, residues with similar chemical behavior have different preferences to be KRs: Arg, Tyr, Val and Thr are preferred over Lys, Trp, Ile and Ser, respectively. Atomic level contacts revealed that charged and polar—nonpolar contacts are dominant in enzymes, polar in structural and nonpolar in regulatory proteins. On the other hand, polar—nonpolar contacts are enriched in all these classes of protein—RNA complexes. Further, the influence of sequence and structural features such as conservation score, surrounding hydrophobicity, solvent accessibility, secondary structure and long-range order in key residues are also discussed. We envisage that the present study provides insights to understand the structural and functional aspects of protein—RNA complexes.

TRNA methyltransferase 5 (Trm5) enzyme is an S-adenosyl methionine (AdoMet)-dependent methyltransferase which methylates the G37 nucleotide at the N¹ atom of the tRNA. The free form of Trm5 enzyme has three intrinsically disordered regions, which are highly flexible and lack stable three-dimensional structures. These regions gain ordered structures upon the complex formation with tRNA, also called disorder-to-order transition (DOT) regions. In this study, we performed molecular dynamics (MD) simulations of archaeal Trm5 in free and complex forms and observed that the DOT residues are highly flexible in free proteins and become stable in complex structures. The energetic contributions show that DOT residues are important for stabilising the complex. The DOT1 and DOT2 are mainly observed to be important for stabilising the complex, while DOT3 is present near the active site to coordinate the interactions between methyl-donating ligands and G37 nucleotides. In addition, mutational studies on the Trm5 complex showed that the wild type is more stable than the G37A tRNA mutant complex. The loss of productive interactions upon G37A mutation drives the AdoMet ligand away from the 37th nucleotide, and Arg145 in DOT3 plays a crucial role in stabilising the ligand, as well as the G37 nucleotide, in the wild-type complex. Further, the overall energetic contribution calculated using MMPBSA corroborates that the wild-type complex has a better affinity between Trm5 and tRNA. Overall, our study reveals that targeting DOT regions for binding could improve the inhibition of Trm5.