Analysis of the interactions observed in four domains of the wt NP and R361A mutant.

Loop 1 (residues 73–90), loop2 (residues 200–211), β-sheet 1 (residues 91–112), and the linker (residues 360–373).These interactions define a path between loop 1 and the residue R361 of the linker, located in the RNA binding groove (see Figures 2 and 4).1HB stands for hydrogen bond.2HPh stands for hydrophobic interaction.See the Experimental section for definitions of the contact domains and the two contact types.a)Loop 1-loop 2 contact: In R361A, hydrophobic interactions between L79 and loop 2 drove loop 1 to contact loop2; transient salt-bridges between R204 or R208, on the one hand, and E80 or E81, on the other hand, stabilized the interaction between loop 1 and loop 2 at short distances. Such loop-loop interactions were not found in wt NP.b)Loop 1 stability (base): The side-chain of K113 was engaged in a strong hydrogen bond with the C-terminus of the loop 1; the guanidinium moiety of K113 formed a salt bridge with E73 at the N-terminus of loop 1, contributing to the stability of loop 1 in wt NP. The R361A mutation drastically reduced the interactions of the K113 with the loop 1, increasing loop 1 flexibility.c)Linker-β sheet 1 contacts: The linker was connected to the β sheet 1 through conserved hydrophobic interaction between M371 and the ring of Y97, stable hydrogen bonds between R106 and the linker backbone oxygen atoms. The R361A mutation decreased the population of the solvated K103-E372 salt-bridge.d)Inter-linker contacts: In NP, E369 interacted with both R361 and R317. In R361A, the R317-E369 contact population increased to 93% as compared to 73% in wt NP and the R361-E369 interaction was canceled by the mutation.