A curated rotamer library for common post-translational modifications of proteins

Abstract

Motivation: Sidechain rotamer libraries of the common amino acids of a protein are useful for folded protein structure determination and for generating ensembles of intrinsically disordered proteins (IDPs). However, much of protein function is modulated beyond the translated sequence through the introduction of post-translational modifications (PTMs).

Results: In this work, we have provided a curated set of side chain rotamers for the most common PTMs derived from the RCSB PDB database, including phosphorylated, methylated, and acetylated sidechains. Our rotamer libraries improve upon existing methods such as SIDEpro, Rosetta, and AlphaFold3 in predicting the experimental structures for PTMs in folded proteins. In addition, we showcase our PTM libraries in full use by generating ensembles with the Monte Carlo Side Chain Entropy (MCSCE) for folded proteins, and combining MCSCE with the Local Disordered Region Sampling algorithms within IDPConformerGenerator for proteins with intrinsically disordered regions.

Availability and implementation: The codes for dihedral angle computations and library creation are available at https://github.com/THGLab/ptm_sc.git.

Graphical Abstract

Figure 1.

Ramachandran plots color coded by χ₁ angle ranges for PTM-modified amino acids using the backbone-dependent library. Backbone bins in 60° separation are shown in gray dash lines. We consider phosphorylated serine (SEP), phosphorylated threonine (TPO), phosphorylated tyrosine (PTR), and tri-methylated lysine (M3L).

Figure 2.

Ramachandran plots color coded by χ₁ and χ₂ angle ranges for SEP and TPO with hydrogen bond formation using the backbone-dependent library. Backbone bins in 60° separation are shown in gray dash lines. Hydrogen bonds are defined by within a donor–acceptor distance cutoff of 3.5 Å. (A1, A2, and B) Representative configurations for SEP (A1, A2) and TPO (B) associated with the annotated backbone regions.

Figure 3.

RMSD distributions of repacked PTM-modified residues using different rotamer libraries and compared to experimental structures and AlphaFold3. SIDEpro does not support ALY packing. (A) Boxplots for the RMSD distributions. Medians are highlighted in white and each box extends from the first quartile (Q1) to the third quartile (Q3). Outliers in circle are defined as points outside of 1.5 times the interquartile range below Q1 or above Q3. (B) Repacked PTM-containing structures using different rotamer libraries compared to the experimental PDB structures. (C) Repacked PTM-containing structures using BD-rotamer library compared to AlphaFold3 and the experimental PDB structures. Examples are taken from PDB ID 3CLY (PTR), 4EZH (M3L), and 4QUT (ALY).

Figure 4.

Histone H3 conformers generated with different PTMs libraries. (A) Modifications on the histone H3 N-terminus on chains C/G of the nucleosome. (B) Ensembles of 30 all-atom H3-modified nucleosome conformers (folded domains and DNA are taken from PDB ID 8SIY). PTM-modified residues are highlighted with stick representations. (C) Comparison of torsion angle probability distributions for PTM-modified residues in the H3 conformers with different libraries. (D) Fractional inter-residue contacts (C_α-C_α distances within 8 Å) of PTMs containing ensembles (δ_ptm) subtracted by the ensemble without modifications (δ₀) for the IDR regions. The maps were calculated with 500 randomly sampled conformers [based on convergence of Rgand averaged over 10 trials. From left to right: BD-rotamer, BI-rotamer, SIDEpro, Rosetta, and Rosetta without clash filtering, with contacts averaged from chain C and G.

Figure 5.

UDF1 conformers generated with different PTM libraries. (A) Modifications on the C-terminal IDR of UDF1 at Y219. (B) Cartoon representations of 30 all-atom UDF1 conformers (structure of the folded domain taken from PDB ID 2YUJ model). (C) Comparison of torsion angle probability distributions for PTM in the UDF1 conformers with different libraries. (D) Fractional inter-residue contacts (C_α–C_α distances within 8 Å) of PTMs containing ensembles (δ_ptm) subtracted by the ensemble without modifications (δ₀) for the IDR region around the PTM site. The maps were calculated with ensembles of 500 randomly sampled conformers and averaged over 10 trials. Top: BD-rotamer, BI-rotamer; bottom (L to R): SIDEpro, Rosetta, and Rosetta without clash filtering.