14-3-3 proteins activate Pseudomonas exotoxins-S and -T by chaperoning a hydrophobic surface

Karlberg, T.; Hornyak, P.; Pinto, A. F.; Milanova, S.; Ebrahimi, M.; Lindberg, M.; Pullen, N.; Nordstrom, A.; Loverli, E.; Caraballo, R.; Wong, E. V.; Nareoja, K.; Thorsell, A.-G.; Elofsson, M.; De La Cruz, E. M.; Bjorkegren, C.; Schuler, H.

doi:10.1101/342659

bioRxiv · 10.1101/342659

14-3-3 proteins activate Pseudomonas exotoxins-S and -T by chaperoning a hydrophobic surface

Karlberg, T.·Hornyak, P.·Pinto, A. F.·Milanova, S.·Ebrahimi, M.·Lindberg, M.·Pullen, N.·Nordstrom, A.·Loverli, E.·Caraballo, R.·Wong, E. V.·Nareoja, K.·Thorsell, A.-G.·Elofsson, M.·De La Cruz, E. M.·Bjorkegren, C.·Schuler, H.

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Abstract

Pseudomonas are a common cause of hospital acquired infections that may be lethal. ADP-ribosyltransferase activities of Pseudomonas exotoxin-S and -T depend on 14-3-3 proteins inside the host cell. By binding in the 14-3-3 phosphopeptide binding groove, a hydrophobic C-terminal helix of ExoS and ExoT has been thought to be crucial for their activation. However, crystal structures of the 14-3-3{beta}:ExoS and -ExoT complexes presented here reveal an extensive novel binding interface that is sufficient for complex formation and toxin activation. We show that C-terminally truncated ExoS ADP-ribosyltransferase domain lacking the hydrophobic binding motif is active when co-expressed with 14-3-3. Moreover, swapping the hydrophobic C-terminus with a fragment from Vibrio Vis toxin creates a 14-3-3 independent toxin that ADP-ribosylates known ExoS targets. Finally, we show that 14-3-3 stabilizes ExoS against thermal aggregation. Together, this indicates that 14-3-3 proteins activate exotoxin ADP-ribosyltransferase domains by chaperoning their hydrophobic surfaces independently of the hydrophobic C-terminal segment.\n\nShort summaryCrystal structures of Pseudomonas exotoxins-S and -T identify a novel hydrophobic interface with 14-3-3 proteins, and we show that 14-3-3 activates these toxins independent of their phosphopeptide groove binding C-termini, by preventing their aggregation.

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Karlberg, T., Hornyak, P., Pinto, A. F., Milanova, S., Ebrahimi, M., Lindberg, M., Pullen, N., Nordstrom, A., Loverli, E., Caraballo, R., Wong, E. V., Nareoja, K., Thorsell, A.-G., Elofsson, M., De La Cruz, E. M., Bjorkegren, C., Schuler, H.. 2018-06-08. 14-3-3 proteins activate Pseudomonas exotoxins-S and -T by chaperoning a hydrophobic surface. https://doi.org/10.1101/342659

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