Biochemical and Structural Consequences of NEDD8 Acetylation

Abstract Similar to ubiquitin, the ubiquitin‐like protein NEDD8 is not only conjugated to other proteins but is itself subject to posttranslational modifications including lysine acetylation. Yet, compared to ubiquitin, only little is known about the biochemical and structural consequences of site‐specific NEDD8 acetylation. Here, we generated site‐specifically mono‐acetylated NEDD8 variants for each known acetylation site by genetic code expansion. We show that, in particular, acetylation of K11 has a negative impact on the usage of NEDD8 by the NEDD8‐conjugating enzymes UBE2M and UBE2F and that this is likely due to electrostatic and steric effects resulting in conformational changes of NEDD8. Finally, we provide evidence that p300 acts as a position‐specific NEDD8 acetyltransferase.