Biology subjects

Listov, D.

Publications and source records attributed to Listov, D..

3 recordsLinked to original sources

bioRxiv2022

The C-terminal tail of CSNAP attenuates the CSN complex

Protein degradation is one of the essential mechanisms that enables reshaping of the proteome landscape in response to various stimuli. The largest E3 ubiquitin ligase family that targets proteins to degradation by catalyzing ubiquitnation is the cullin-RING ligases (CRL). Many of the proteins that are regulated by CRLs are central to tumorigenesis and tumour progression, and dysregulation of the CRL family is frequently associated with cancer. The CRL family comprises [~]300 complexes all of which are regulated by the COP9 signalosome complex (CSN). Therefore, the CSN is considered an attractive target for therapeutic intervention. Research efforts for targeted CSN inhibition have been directed towards inhibition of the complex enzymatic subunit, CSN5. Here, we have taken a fresh approach focusing on CSNAP, the smallest CSN subunit. Our results show that the C-terminal region of CSNAP is tightly packed within the CSN complex, in a groove formed by CSN3 and CSN8. We show that a 16 amino acid C-terminal peptide, derived from this CSN interacting region, can displace the endogenous CSNAP subunit from the complex. This, in turn, leads to a CSNAP null phenotype that attenuates CSN activity and consequently CRLs function. Overall, our findings emphasize the potential of a CSNAP-based peptide for CSN inhibition as a new therapeutic avenue.

biochemistry↗

bioRxiv2022

Protein Quaternary Structures in Solution are a Mixture of Multiple forms

Over half the proteins form homo or hetero-oligomeric structures. Experimentally determined structures are often considered in determining a proteins oligomeric state, but static structures miss the dynamic equilibrium between different quaternary forms. The problem is exacerbated in homo-oligomers, where the oligomeric states are challenging to characterize. Here, we re-evaluated the oligomeric state of 17 different bacterial proteins across a broad range of protein concentrations and solutions by native mass-spectrometry (MS), mass photometry (MP), size exclusion chromatography (SEC), and small-angle x-ray scattering (SAXS), finding that most exhibit several oligomeric states. Surprisingly, many proteins did not show mass-action driven equilibrium between the oligomeric states. For approximately half the proteins, the predicted oligomeric forms described in publicly available databases underestimated the complexity of protein quaternary structures in solution. Conversely, AlphaFold Multimer provided an accurate description of the potential multimeric states for most proteins, suggesting that it could help resolve uncertainties on the solution state of many proteins.

biochemistry↗

bioRxiv2021

Assessing and enhancing foldability in designed proteins

Recent advances in protein-design methodology have led to a dramatic increase in reliability and scale. With these advances, dozens and even thousands of designed proteins are automatically generated and screened. Nevertheless, the success rate, particularly in design of functional proteins, is low and fundamental goals such as reliable de novo design of efficient enzymes remain beyond reach. Experimental analyses have consistently indicated that a major reason for design failure is inaccuracy and misfolding relative to the design model. To address this challenge, we describe complementary methods to diagnose and ameliorate suboptimal regions in designed proteins: first, we develop a Rosetta atomistic computational mutation scanning approach to detect energetically suboptimal positions in designs (available on a web server https://pSUFER.weizmann.ac.il); second, we demonstrate that AlphaFold2 ab initio structure prediction flags regions that may misfold in designed enzymes and binders; and third, we focus FuncLib design calculations on suboptimal positions in a previously designed low-efficiency enzyme improving its catalytic efficiency by 330 fold. Furthermore, applied to a de novo designed protein that exhibited limited stability, the same approach markedly improved stability and expressibility. Thus, foldability analysis and enhancement may dramatically increase the success rate in design of functional proteins.

biochemistry↗