Illuminating the dark protein-protein interactome

Tabar, Mehdi Sharifi; Parsania, Chirag; Chen, Hong; Bailey, Charles G.; Rasko, John E.J.

doi:10.1016/j.crmeth.2022.100275

Cited by 22 publications

(15 citation statements)

References 77 publications

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“…Proteins are the workhorses of the cell, performing the vast majority of cellular functions, much of which occur in the context of higher order protein complexes. Consequently, there is a broad interest to better understand the largely hidden 'social' nature of proteins, especially those serving as hubs within complex regulatory networks (20)(21)(22)(23)(24)(25). In this study, we investigated the interactome of MecA due to its broad conservation, potential role as a major network hub, and the limited number of known MecA interactions.…”

Section: Discussionmentioning

confidence: 99%

“…Recent improvements in the technologies used to detect and characterize protein-protein interactions (PPI) have stimulated new interest to reveal protein interactomes, leading to the discovery of many unexpected PPI, especially those involved in previously unrecognized moonlighting functions (20)(21)(22)(23)(24)(25). Several genetic and biochemical techniques are commonly employed to detect PPI, such as two-hybrid assays, coimmunoprecipitation (co-IP), bimolecular fluorescence complementation (BiFC), and Förster resonance energy transfer (FRET) (26)(27)(28)(29).…”

Section: Introductionmentioning

confidence: 99%

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Mass spectrometry and split luciferase complementation assays reveal the MecA protein interactome ofStreptococcus mutans

Qin,

Anderson,

Zou

et al. 2023

Preprint

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MecA is a highly conserved adaptor protein encoded by prokaryotes from theBacillotaphylum. MecA mutants exhibit similar pleiotropic defects in a variety of organisms, although most of these phenotypes currently lack a mechanistic basis. MecA mediates ClpCP-dependent proteolysis of its substrates, but only several such substrates have been reported in the literature and there are suggestions that proteolysis-independent regulatory mechanisms may also exist. Here, we provide the first comprehensive characterization of the MecA interactome and further assess its regulatory role in Clp-dependent proteolysis. Untargeted coimmunoprecipitation assays coupled with mass spectrometry revealed that the MecA ortholog from the oral pathobiontStreptococcus mutanslikely serves as a major protein interaction network hub by potentially complexing with >100 distinct protein substrates, most of which function in highly conserved metabolic pathways. The interactome results were independently verified using a newly developed prokaryotic split luciferase complementation assay (SLCA) to detect MecA protein-protein interactionsin vivo. In addition, we further develop a new application of SLCA to supportin vivomeasurements of MecA relative protein binding affinities. SLCA results were independently verified using targeted coimmunoprecipitation assays, suggesting the general utility of this approach for prokaryotic protein-protein interaction studies. Our results indicate that MecA indeed regulates its interactome through both Clp-dependent proteolysis as well as through an as yet undefined proteolysis-independent mechanism that may affect more than half of its protein interactome. This suggests a significant aspect of MecA regulatory function still has yet to be discovered.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Mass spectrometry and split luciferase complementation assays reveal the MecA protein interactome ofStreptococcus mutans

Qin,

Anderson,

Zou

et al. 2023

Preprint

View full text Add to dashboard Cite

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“…[ 7 ] In part, the apparently poor reproducibility has a clear biological background. [ 30 ] For example, as mentioned above, 2‐hybrid qualitatively studies intrinsic interactomes, but AP‐MS qualitatively studies cellular complexomes. Consequently, 2‐hybrid identifies pairs of molecules that could directly interact, while AP‐MS identifies complexes that may contain indirectly bound components.…”

Section: The Issue Of Reproducibility In Qualitative Interactomicsmentioning

confidence: 99%

Molecules interact. But how strong and how much?

2023

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Interactomics aims to characterize all interactions formed between molecules that comprise our body. Although it emerged from quantitative biophysics, it has devolved into a predominantly qualitative field of science over the past decades. Due to technical limitations at its onset, almost all tools in interactomics are qualitative, which persists in defining the discipline. Here, we argue that interactomics needs to return to a quantitative direction because the technical achievements of the last decade have overcome the original limitations that forced its current path. In contrast to qualitative interactomics which is constrained to charting lists of observed interactions, quantitative interactomics can also uncover answers to key questions such as the strength of interactions or how many of certain complexes can form in cells, thus providing researchers with more immediate proxies for understanding and predicting biological processes.

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“…The human molecular interaction system is the one that underlies the human body. It is estimated that there are at least 140,000 pairwise protein–protein interactions (PPIs) 76 . In order to modulate the downstream signaling events, intensive research has been conducted on innovative peptides to counteract pathogenic PPIs.…”

Section: Interactions Between Proteins and Peptidesmentioning

confidence: 99%

“…It is estimated that there are at least 140,000 pairwise protein-protein interactions (PPIs). 76 In order to modulate the downstream signaling events, intensive research has been conducted on innovative peptides to counteract pathogenic PPIs. The large surface area of the larger PPI interfaces makes it difficult to modulate the small molecules (generally 1500-3000 Å2), in contrast to the binding pockets of the small molecules (300-1000 Å2).…”

Section: Interactions Between Proteins and Peptidesmentioning

confidence: 99%

Translational aspect in peptide drug discovery and development: An emerging therapeutic candidate

et al. 2022

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In the last two decades, protein-protein interactions (PPIs) have been used as the main target for drug development. However, with larger or superficial binding sites, it has been extremely difficult to disrupt PPIs with small molecules. On the other hand, intracellular PPIs cannot be targeted by antibodies that cannot penetrate the cell membrane. Peptides that have a combination of conformational rigidity and flexibility can be used to target difficult binding interfaces with appropriate binding affinity and specificity. Since the introduction of insulin nearly a century ago, more than 80 peptide drugs have been approved to treat a variety of diseases. These include deadly diseases such as cancer and human immunodeficiency virus infection. It is also useful against diabetes, chronic pain, and osteoporosis. Today, more research is being done on these drugs as lessons learned from earlier approaches, which are still valid today, complement newer approaches such as peptide display libraries. At the same time, integrated genomics and peptide display libraries are new strategies that open new avenues for peptide drug discovery. The purpose of this review is to examine the problems in elucidating the peptide-protein recognition mechanism. This is important to develop peptide-based interventions that interfere with endogenous protein interactions. New approaches are being

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Illuminating the dark protein-protein interactome

Cited by 22 publications

References 77 publications

Mass spectrometry and split luciferase complementation assays reveal the MecA protein interactome ofStreptococcus mutans

Mass spectrometry and split luciferase complementation assays reveal the MecA protein interactome ofStreptococcus mutans

Molecules interact. But how strong and how much?

Translational aspect in peptide drug discovery and development: An emerging therapeutic candidate

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