The effect of N-acetylation and N-methylation of lysine residue of Tat peptide on its interaction with HIV-1 TAR RNA.

Santosh Kumar,Souvik Maiti,Fatah Kashanchi

doi:10.1371/journal.pone.0077595

Santosh Kumar, Souvik Maiti + Show 1 more

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https://doi.org/10.1371/journal.pone.0077595

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Abstract

Post-translational modification (PTM) of RNA binding proteins (RBPs) play a very important role in determining their binding to cognate RNAs and therefore regulate the downstream effects. Lysine can undergo various PTMs and thereby contribute to the regulation of different cellular processes. It can be reversibly acetylated and methylated using a pool of respective enzymes, to act as a switch for controlling the binding efficiency of RBPs. Here we have delineated the thermodynamic and kinetic effects of N-acetylation and N-monomethylation of lysine on interaction between HIV-1 TAR RNA and its cognate binder Tat peptide ( a model system). Our results indicate that acetylation of lysine 50 (K50), leads to eight- fold reduction in binding affinity, originating exclusively from entropy changes whereas, lysine 51 (K51) acetylation resulted only in three fold decrease with large enthalpy-entropy compensation. The measurement of kinetic parameters indicated major change (4.5 fold) in dissociation rate in case of K50 acetylation however, K51 acetylation showed similar effect on both association and dissociation rates. In contrast, lysine methylation did not affect the binding affinity of Tat peptide to TAR RNA at K50, nonetheless three fold enhancement in binding affinity was observed at K51 position. In spite of large enthalpy-entropy compensation, lysine methylation seems to have more pronounced position specific effect on the kinetic parameters. In case of K50 methylation, simultaneous increase was observed in the rate of association and dissociation leaving binding affinity unaffected. The increased binding affinity for methylated Tat at K51 stems from faster association rate with slightly slower dissociation rate.

Highlights

Post-translational modifications (PTMs) of RNA binding proteins (RBPs) regulate many important cellular functions including nuclear transport, various signaling pathways and gene expression during stress conditions [1,2,3]
Thermodynamic parameters corresponding to Tat-transactivation region (TAR) RNA binding along with their binding affinity (KA) and stoichiometry (n) of binding are listed in the Table 1
Our results provide insight into the molecular understanding of how acetylation of lysine at K50 position leads to the inability of Tat peptide binding to TAR RNA [31], on the other hand methylation, another type of PTM of lysine at K51 position results in transcriptional activation [41]

Summary

Introduction

Post-translational modifications (PTMs) of RNA binding proteins (RBPs) regulate many important cellular functions including nuclear transport, various signaling pathways and gene expression during stress conditions [1,2,3]. Most of the RBPs are rich in positively charged amino acids which mediate the initial recognition of the negatively charged phosphate backbone of the RNA followed by the formation of specific Hbonds mediated by side chain, terminal protons of amino and guanidinium groups respectively for lysine and arginine [4,5]. Any alteration in this molecular arrangement would result in a change in binding strength of RNA and protein. Lysine methylation has been linked to a plethora of disease conditions related to growth and development of the organism, where levels of these enzymes are altered [14]

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