Protein Design Part II

Part A: SOD1 Binder Peptide Design (From Pranam)

What I know about SOD1 and its mutation:

Mutations in SOD1 cause familial Amyotrophic Lateral Sclerosis (ALS)
ALS is a heterogeneous severe neurodegenerative disorder the hallmark of which is an adult-onset loss of upper and lower motor neurons.
It leads to a progressive paresis and atrophy of skeletal muscles, resulting in quadriplegia and fatal respiratory failure.
The mutation subtly destabilizes the N-terminus, perturbs folding energetics, and promotes toxic aggregation.

Challenge of this week: Design short peptides that bind mutant SOD1 & then decide which ones are worth advancing toward therapy.

Part 1A: Generate Binders with PepMLM

To generate binders using the suggested program, it’s necessary to have the original sequence and check for the A4V mutation at that position.

Original sequence Uniprot (P00441)

MATKAVCVLKGDGPVQGIINFEQKESNGPVKVWGSIKGLTEGLHGFHVHEFGDNTAGCTSAGPHFNPLSRKHGGPKDEERHVGDLGNVTADKDGVADVSIEDSVISLSGDHCIIGRTLVVHEKADDLGKGGNEESTKTGNAGSRLACGVIGIAQ

A4V Mutation sequence:

MATKVVCVLKGDGPVQGIINFEQKESNGPVKVWGSIKGLTEGLHGFHVHEFGDNTAGCTSAGPHFNPLSRKHGGPKDEERHVGDLGNVTADKDGVADVSIEDSVISLSGDHCIIGRTLVVHEKADDLGKGGNEESTKTGNAGSRLACGVIGIAQ

After having the sequence modified, we use the Colab notebook: Important to know that it’s to make sure you select the number 4 of binders in the input, select the length of peptides, and then run it

Input

Binders and Peptide Length

Table 1. Peptides predicted

Index	Length	Binder	Pseudo Perplexity (score)*
Pep 0	12	WRYPAVGARWKX	10.660527
Pep 1	12	WRYPVAAVELKX	10.027294
Pep 2	12	WLYYPAGAAHWX	11.046032
Pep 3	12	KRSYVVGVEWGX	17.759518
Control**	12	FLYRWLPSRRGG	——————

Description: (*) Pseudo perplexity is an adaptation of the perplexity metric used in masked language models. The model masks each amino acid in the peptide one at a time and estimates the probability of correctly recovering it given the surrounding residues and the target protein sequence. Lower value → model assigns a higher probability to the peptide sequence and high confidence. High value → Less confidence model of sequence for the peptide. (**) Control is a known SOD1-binding peptide.

Based on the results in Table 1, the candidates are in the top 2 positions. And less confidence with the last position (index 3).

Part 2A: Evaluate Binders with AlphaFold3

To evaluate the generated binders, AlphaFold3 was used to model protein–peptide complexes.