Effect of D122N mutation in BAK1 on structural integrity of protein complex consisting FLS2, flg22 and BAK1 ectodomain- a molecular dynamics study”

Abstract

Elongated (elg) phenotype of Arabidopsis thaliana is caused by D122N mutation in the third leucine repeat of BRI1-associated kinase 1 (BAK1) protein that changes the 122nd amino acid aspartate to asparagine. The mutation is mainly associated with photomorphogenesis since plants with this mutation are characterized to be early flowering and have elongated stem and petioles. However, the BAK1 protein also plays a crucial role in plant immunity by forming a heterodimer with pattern triggered receptor Flagellin sensing 2 (FLS2) upon the perception of flg22, an elicitor protein from bacterial flagellin, that cause transphosphorylation of BAK1 and FLS2 and subsequent initiation of signal transduction pathway leading to activation of the immune response. Here we investigated the impact of D122N mutation in BAK1 on the structural integrity of the FLS2-flg22-BAK1 complex through molecular dynamics simulation. We induced the D122N mutation in BAK1 protein of native FLS2-flg22-BAK1 crystallographic structure using the in-silico method. Subsequently, we simulated both native and mutated complex using molecular dynamics methods for 30ns. The simulations were compared using parameters such as root mean square deviation (RMSF), root mean square fluctuation (RMSF), the radius of gyration (Rg) and the number of hydrogen bonds to compared the structural integrity of two complexes. We found that the mutation resulted in often 2 Armstrong higher deviation of atoms from reference structure for the whole complex, FLS2, BAK1, and flg22 compared to counterpart from the non-mutated complex. RMSF analysis revealed that mutation caused higher fluctuation of amino acid in Both the N terminal and C terminal of FLS2 that hinder interaction with both flg22 and BAK1. Except for flg22, both FLS2 and BAK1 had a higher radius of gyration in the complex containing mutation compared to the non-mutated complex. The mutation hindered the formation of hydrogen bonds among all the peptides, often terminating all the hydrogen bonds between peptide. Even though ASP122 does not form a direct connection with FLS2 atoms, the mutation results in the termination of intra-protein interaction of 122nd amino acid with ARG146 in BAK1, which undermines the interaction of BAK1 peptide with FLS2.