Phosphorylation of the conserved C‐terminal domain of ribosomal P‐proteins impairs the mode of interaction with plant toxins

Abstract

The ribosome is subjected to post‐translational modifications, including phosphorylation, that affect its biological activity. Among ribosomal elements, the P‐proteins undergo phosphorylation within the C terminus, the element which interacts with trGTPases or ribosome‐inactivating proteins (RIPs); however, the role of phosphorylation has never been elucidated. Here, we probed the function of phosphorylation on the interaction of P‐proteins with RIPs using the ribosomal P1‐P2 dimer. We determined the kinetic parameters of the interaction with the toxins using biolayer interferometry and microscale thermophoresis. The results present the first mechanistic insight into the function of P‐protein phosphorylation, showing that introduction of a negative charge into the C terminus of P1‐P2 proteins promotes α‐helix formation and decreases the affinity of the P‐proteins for the RIPs.

Cryo-EM structure of Shiga toxin 2 in complex with the native ribosomal P-stalk reveals residues involved in the binding interaction

“…Interestingly, fitting the density with a short α-helix satisfies the hydrogen bonding pattern described previously. The observation that the CTD might behave as a random coil or as an α-helix is of significance, and it indicates the possibility for structural rearrangements in this region, as indicated previously ( 28 ).…”

“…This finding implicates a random coil to an α-helix transition in the CTD upon binding to Stx2a. This observation is supported by the molecular dynamics simulation of the TCS–CTD interaction, which indicated that the CTD has the propensity for α-helix formation in the presence of TCS ( 28 ).

Figure 8 The binding interfaces formed by the C-terminal peptides from the CTDs of P-proteins with RIPs and trGTPases.

…”

Single-domain antibodies neutralize ricin toxin intracellularly by blocking access to ribosomal P-stalk proteins

“…The catalytic center of RTA constitutes a solvent-exposed cleft on one side of the molecule that accommodates a protruding adenine base. While the molecular basis of the depurination reaction has been recognized for decades, only recently have we gained insight into how RTA is recruited to the ribosome in the first place ( 19 , 24 , 25 , 39 , 40 , 41 ). Indeed, there is now compelling evidence that the RIP activity of RTA is actually a two-step event, wherein step 1 involves interactions with ribosome by P-stalk proteins before the second engagement with the SRL.…”

A Sixty-Year Research and Development of Trichosanthin, a Ribosome-Inactivating Protein

“…In 2017, Grela et al showed functional non-equivalence of the individual P-protein CTDs [ 69 ]. In 2021, Horbowicz-Drożdżal et al revealed that phosphorylation of the conserved C-terminal domain of ribosomal P-proteins impairs the interaction with plant toxins [ 70 ]. The study of how TCS interacts with ribosomes became a model for similar work in other RIPs [ 71 ].…”