Evaluation of affinity-purification coupled to mass spectrometry approaches for capture of short linear motif-based interactions

Kassa, Eszter; Jamshidi, Sara; Mihalič, Filip; Simonetti, Leandro; Kliche, Johanna; Jemth, Per; Lind, Sara Bergström; Ivarsson, Ylva

doi:10.1016/j.ab.2022.115017

Cited by 11 publications

(6 citation statements)

References 49 publications

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“…Here, we developed an innovative top-down quantitative interactomic strategy that combines two state-of-the-art holdup-based methods developed by our team. Interactions mediated by short linear motifs are most often highly transient and routinely used interactomic techniques often fail to detect them (Kassa et al , 2023). Our new strategy was found to be particularly powerful to identify and characterize these interactions.…”

Section: Discussionmentioning

confidence: 99%

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Uncovering the BIN1-SH3 interactome underpinning centronuclear myopathy

Zámbó

Edelweiss

Morlet

et al. 2023

Preprint

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Deletion of the protein-protein interaction SH3 domain of the membrane remodeling BIN1 protein was found to lead to centronuclear myopathy in patients, yet only few interaction partners of BIN1 SH3 have been identified so far. Here we used the holdup assay to proteome-wide measure steady-state affinity constants of BIN1 SH3 domain for thousands of full-length cellular proteins, as well as for hundreds of putative SH3-binding sites found within the identified BIN1 binders. Besides confirming known partners, such as DNM2, we also identified and affinity-characterized numerous others. We also assessed the impact of a set of rare natural BIN1 SH3 domain variants on affinity interactomes and identified a set of potentially harmful ones that exhibited perturbed affinity profiles, whose impacts were confirmed by cellular assay, tentatively connecting them to neuromuscular disorders.

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Section: Discussionmentioning

confidence: 99%

“…Interactions mediated by short linear motifs are highly transient and routinely used interactomic techniques often fail to detect them (Kassa et al, 2023). Consequently, past studies only identified a handful of BIN1 interaction partners (Figure 1).…”

Section: Discussionmentioning

confidence: 99%

Uncovering the BIN1-SH3 interactome underpinning centronuclear myopathy

Zámbó

Edelweiss

Morlet

et al. 2023

Preprint

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“…Through testing eight distinct peptide sequences with varying affinities for three specific protein domains (KEAP1 Kelch, MDM2 SWIB, and TSG101 UEV), the study revealed that biotin-peptide pulldown outperformed PRISMA in validating SLiMs. Notably, tandem peptide repeats enhanced interaction capture, underscoring the need to consider method development parameters for effective affinity capture MS-based validation of SLiM-based interactions from cell lysates [20]. Moreover, a new method, thermal proximity coaggregation (TPCA), has been introduced to monitor the dynamics of native protein complexes in living cells.…”

Section: High-throughput Ppi Detection Methods and Dmismentioning

confidence: 99%

“…Alternative exons exhibit a notable enrichment of IDRs, showing their importance in functional diversity [17,18]. Interactions facilitated by SLiMs are transient and exhibit low affinity, typically within 1-150 lM [19,20]. It is important to note that SLiMs usually comprise only 2-5 precisely defined positions, posing a challenge for their identification using experimental and computational approaches [21].…”

mentioning

confidence: 99%

Uncovering domain motif interactions using high‐throughput protein–protein interaction detection methods

Idrees,

Paudel,

Sadaf

et al. 2024

FEBS Letters

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Protein–protein interactions (PPIs) are often mediated by short linear motifs (SLiMs) in one protein and domain in another, known as domain–motif interactions (DMIs). During the past decade, SLiMs have been studied to find their role in cellular functions such as post‐translational modifications, regulatory processes, protein scaffolding, cell cycle progression, cell adhesion, cell signalling and substrate selection for proteasomal degradation. This review provides a comprehensive overview of the current PPI detection techniques and resources, focusing on their relevance to capturing interactions mediated by SLiMs. We also address the challenges associated with capturing DMIs. Moreover, a case study analysing the BioGrid database as a source of DMI prediction revealed significant known DMI enrichment in different PPI detection methods. Overall, it can be said that current high‐throughput PPI detection methods can be a reliable source for predicting DMIs.

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“…In combination with the parallel synthesis of peptides on cellulose membranes (SPOT synthesis) 25 , the throughput of peptide-based interaction proteomics can be greatly increased 24,26,27 . This Protein Interaction Screen on Peptide Matrix (PRISMA) method has been applied in a number of recent studies [28][29][30] . For example, we used PRISMA to study how pathogenic mutations in IDRs affect proteinprotein interactions (PPIs).…”

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confidence: 99%

Pathogenic mutations of human phosphorylation sites affect protein-protein interactions

Rrustemi

Meyer

Roske

et al. 2023

Preprint

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Despite their lack of a defined 3D structure, intrinsically disordered regions (IDRs) of proteins play important biological roles. Many IDRs contain short linear motifs (SLiMs) that mediate protein-protein interactions (PPIs), which can be regulated by post-translational modifications like phosphorylation. 20% of pathogenic missense mutations are found in IDRs, and understanding how such mutations affect PPIs is essential for unraveling disease mechanisms. Here, we employed peptide-based interaction proteomics to investigate 36 disease-causing mutations affecting phosphorylation sites. Our results unveiled significant differences in interactomes between phosphorylated and non-phosphorylated peptides, often due to disrupted phosphorylation-dependent SLiMs. We focused on a mutation of a serine phosphorylation site in the transcription factor GATAD1, which causes dilated cardiomyopathy. We found that this phosphorylation site mediates interaction with 14-3-3 family proteins. Follow-up experiments revealed the structural basis of this interaction and suggest that 14-3-3 binding affects GATAD1 nucleocytoplasmic transport by masking a nuclear localisation signal. Our results demonstrate that pathogenic mutations of human phosphorylation sites can significantly impact protein-protein interactions, offering fresh insights into potential molecular mechanisms underlying pathogenesis.

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Evaluation of affinity-purification coupled to mass spectrometry approaches for capture of short linear motif-based interactions

Cited by 11 publications

References 49 publications

Uncovering the BIN1-SH3 interactome underpinning centronuclear myopathy

Uncovering the BIN1-SH3 interactome underpinning centronuclear myopathy

Uncovering domain motif interactions using high‐throughput protein–protein interaction detection methods

Pathogenic mutations of human phosphorylation sites affect protein-protein interactions

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