Biology subjects

Suhr, C. H.

Publications and source records attributed to Suhr, C. H..

2 recordsLinked to original sources

bioRxiv2025

Blocking protein quality control degradation leads to structural stabilization of DHFR indel variants

Gene variants leading to insertions or deletions of amino acid residues (indels) often have detrimental consequences for the folding of the encoded protein. Yet at some positions indels are tolerated or result merely in partial unfolding and hypomorphic phenotypes. Typically unfolded proteins are targeted for protein quality control (PQC) degradation via the ubiquitin-proteasome system, which in yeast is mediated by specific E3 ubiquitin-protein ligases, including Ubr1 and San1. Here we systematically probed the folding of a library of indel variants in the DHFR protein using a sensitive yeast-based protein folding reporter. We show that deletion of Ubr1 or San1 leads to a greater fraction of folded DHFR indel variants, primarily positioned towards the N- and C-terminal regions in DHFR. Intriguingly, most of the DHFR indels that are structurally stabilized in the E3 knockout strains, are also stabilized at lowered temperatures and upon binding the DHFR inhibitor methotrexate. This suggests that blocking PQC degradation can restore function to partially unfolded hypomorph variants, thus providing a potential therapeutic avenue for protein misfolding diseases.

biochemistry↗

bioRxiv2024

Systematic characterization of indel variants using a yeast-based protein folding sensor

Gene variants resulting in insertions or deletions of amino acid residues (indels) have important consequences for evolution and are often linked to disease, yet compared to missense variants the effects of indels are poorly understood and predicted. To approach this issue, we developed a sensitive protein folding sensor based on complementation of uracil auxotrophy in yeast by circular permutated orotate phosphoribosyltransferase (CPOP). The sensor accurately reports on the folding of disease-linked missense variants and de novo designed proteins. Applying the folding sensor to a saturated library of single amino acid indel variants in human DHFR revealed that most regions which tolerate indels are confined to internal loops and the N- and C-termini. Surprisingly, indels are also allowed at a central -helix. Several indels are temperature-sensitive and the folding of most of these indels is rescued upon binding to the competitive DHFR inhibitor methotrexate. Rosetta and AlphaFold2 predictions correlate with the observed effects, suggesting that most indels operate by destabilizing the native fold and that these computational tools may be useful for classification of indels observed in population sequencing.

biochemistry↗