Residues neighboring an SH3-binding motif participate in the interaction in vivo

In signaling networks, protein-protein interactions are often mediated by modular domains that bind short linear motifs. The motifs’ sequences affect many factors, among them affinity and specificity, or the ability to bind strongly and to the appropriate partners. Using Deep Mutational Scanning to create a mutant library, and protein complementation assays to measure protein-protein interactions, we determined the in vivo binding strength of a library of mutants of a binding motif on the MAP kinase kinase Pbs2, which binds the SH3 domain of the osmosensor protein Sho1 in Saccharomyces cerevisiae. These measurements were made using the full-length endogenous proteins, in their native cellular environment. We find that along with residues within the canonical motif, many mutations in the residues neighboring the motif also modulate binding strength. Interestingly, all Pbs2 mutations which increase affinity are situated outside of the Pbs2 region that interacts with the canonical SH3 bind..., Multiple methods were used for data collection:
DHFR-Protein Complementation Assay in a pooled competition assay
DHFR-Protein Complementation Assay in individual colonies
Growth curves
Protein structure predictions using AlphaFold-Multimer
In Revised Upload:
Sequence and structure alignments using mafft and MUSTANG respectively, , # Data from: **Residues neighboring an SH3-binding motif participate in the interaction *In Vivo***

[https://doi.org/10.5061/dryad.79cnp5j3z](https://doi.org/10.5061/dryad.79cnp5j3z)

## Description of the data and file structure

All data pertaining to the manuscript \"**Residues neighboring an SH3-binding motif participate in the interaction *In Vivo***\" by Jordan et al. 

These datasets can be used with the scripts hosted at [https://github.com/Landrylab/Jordan_et_al_2024](https://github.com/Landrylab/Jordan_et_al_2024]) to recreate the analysis used in the manuscript and to recreate the figures in the manuscript.

Readme.md files in the different folders describe the datasets within, including describing the columns in tabular formatted data.

This repository is designed to be used with the scripts in the accompanying github repo. The unzipped \"Data\" folder should be placed in the same folder as the \"figures\" and \"scripts\" folders from the github repo.

The data is divided into 6 ma...,