Structural basis of Naa20 activity towards a canonical NatB substrate

Dominik Layer,Karine Lapouge,Maja Köhn,Jürgen Kopp,Miriam Fontanillo,Irmgard Sinning

doi:10.1038/s42003-020-01546-4

Abstract

N-terminal acetylation is one of the most common protein modifications in eukaryotes and is carried out by N-terminal acetyltransferases (NATs). It plays important roles in protein homeostasis, localization, and interactions and is linked to various human diseases. NatB, one of the major co-translationally active NATs, is composed of the catalytic subunit Naa20 and the auxiliary subunit Naa25, and acetylates about 20% of the proteome. Here we show that NatB substrate specificity and catalytic mechanism are conserved among eukaryotes, and that Naa20 alone is able to acetylate NatB substrates in vitro. We show that Naa25 increases the Naa20 substrate affinity, and identify residues important for peptide binding and acetylation activity. We present the first Naa20 crystal structure in complex with the competitive inhibitor CoA-Ac-MDEL. Our findings demonstrate how Naa20 binds its substrates in the absence of Naa25 and support prospective endeavors to derive specific NAT inhibitors for drug development.

Highlights

N-terminal acetylation is one of the most common protein modifications in eukaryotes and is carried out by N-terminal acetyltransferases (NATs)
In order to dissect the molecular mechanism of NatB and its use as a potential therapeutic target, we studied Chaetomium thermophilum NatB complex and its individual subunits
Our results show that CtNaa[20] is active towards a canonical NatB substrate without CtNaa[25] in vitro, less efficient than in

Summary

Introduction

N-terminal acetylation is one of the most common protein modifications in eukaryotes and is carried out by N-terminal acetyltransferases (NATs) It plays important roles in protein homeostasis, localization, and interactions and is linked to various human diseases. Terminal acetylation, an acetyl group is transferred from acetyl coenzyme A (AcCoA) to the α-amino group of a polypeptide This modification is mostly carried out co-translationally[7] by Nα-acetyltransferase complexes (NATs), which differ in subunit composition and substrate specificity, but comprise at least one catalytic subunit[1,8]. Naa[20] and the auxiliary subunit Naa[25] (formerly known as Nat[3] and Mdm[20], respectively)[10] Both subunits are conserved within eukaryotic model organisms and the NatB complex is found associated with the ribosome[11,12,13]. Naa[25] was shown to be essential for the activity of Naa2015,16,24 and localizes in the cytoplasm, while Naa[20] is found in both, the nucleus[13] and cytoplasm independent of Naa2511,13

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