Promiscuity mapping of the S100 protein family using a high-throughput holdup assay

Simon, Márton A; Bartus, Éva; Mag, Beáta; Boros, Eszter; Roszjár, Lea; Gógl, Gergő; Travé, Gilles; Martinek, Tamás A.; Nyitray, László

doi:10.1038/s41598-022-09574-2

Cited by 10 publications

(19 citation statements)

References 35 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…S100P is considered as one of the most promiscuous members of the S100 protein family, able to bind numerous ligands with a high affinity, and sharing interaction partners with other S100 proteins [ 60 , 61 ]. Therefore, one may expect that other representatives of the S100 protein family are cross-reactive with the same four-helical cytokines, as previously shown for IL-6 family [ 21 ].…”

Section: Discussionmentioning

confidence: 99%

Calcium-Bound S100P Protein Is a Promiscuous Binding Partner of the Four-Helical Cytokines

Kazakov

Deryusheva

Permyakova

et al. 2022

IJMS

View full text Add to dashboard Cite

S100 proteins are multifunctional calcium-binding proteins of vertebrates that act intracellularly, extracellularly, or both, and are engaged in the progression of many socially significant diseases. Their extracellular action is typically mediated by the recognition of specific receptor proteins. Recent studies indicate the ability of some S100 proteins to affect cytokine signaling through direct interaction with cytokines. S100P was shown to be the S100 protein most actively involved in interactions with some four-helical cytokines. To assess the selectivity of the S100P protein binding to four-helical cytokines, we have probed the interaction of Ca2+-bound recombinant human S100P with a panel of 32 four-helical human cytokines covering all structural families of this fold, using surface plasmon resonance spectroscopy. A total of 22 cytokines from all families of four-helical cytokines are S100P binders with the equilibrium dissociation constants, Kd, ranging from 1 nM to 3 µM (below the Kd value for the S100P complex with the V domain of its conventional receptor, receptor for advanced glycation end products, RAGE). Molecular docking and mutagenesis studies revealed the presence in the S100P molecule of a cytokine-binding site, which overlaps with the RAGE-binding site. Since S100 binding to four-helical cytokines inhibits their signaling in some cases, the revealed ability of the S100P protein to interact with ca. 71% of the four-helical cytokines indicates that S100P may serve as a poorly selective inhibitor of their action.

show abstract

Section: Discussionmentioning

confidence: 99%

Calcium-Bound S100P Protein Is a Promiscuous Binding Partner of the Four-Helical Cytokines

Kazakov

Deryusheva

Permyakova

et al. 2022

IJMS

View full text Add to dashboard Cite

show abstract

“…S100P is considered as one of the most promiscuous members of S100 protein family, able to bind numerous ligands with a high affinity, and sharing interaction partners with other S100 proteins [60,61]. Therefore, one may expect that other representatives of the S100 protein family are cross-reactive with the same four-helical cytokines, as previously shown for IL-6 family [21].…”

Section: Discussionmentioning

confidence: 86%

Calcium-Bound S100P Protein is a Promiscuous Binding Partner of the Four-Helical Cytokines

Kazakov¹,

Deryusheva²,

Permyakova³

et al. 2022

Preprint

View full text Add to dashboard Cite

S100 proteins are multifunctional calcium-binding proteins of vertebrates that act intracellularly, extracellularly, or both, and are engaged in the progression of many socially significant diseases. Their extracellular action is typically mediated by the recognition of specific receptor proteins. Besides, recent studies indicate the ability of some S100 proteins to affect cytokine signaling through direct interaction with cytokines. S100P was shown to be the S100 protein most actively involved in interactions with some of four-helical cytokines. To assess selectivity of S100P protein binding to four-helical cytokines, we have probed interaction of Ca2+-bound recombinant human S100P with a panel of 32 four-helical human cytokines covering all structural families of this fold, using surface plasmon resonance spectroscopy. 22 cytokines from all families of four-helical cytokines are S100P binders with the equilibrium dissociation constants, Kd, ranging from 1 nM to 3 µM (below the Kd value for the S100P complex with the V domain of its conventional receptor, receptor for advanced glycation end products, RAGE). Molecular docking and mutagenesis studies revealed the presence in the S100P molecule of a cytokine-binding site, which overlaps with the RAGE-binding site. Since S100 binding to four-helical cytokines inhibits their signaling in some cases, the revealed ability of S100P protein to interact with ca 71% of the four-helical cytokines indicates that S100P may serve as a poorly selective inhibitor of their action.

show abstract

“…In nHU, total cell extracts are used as an analyte, and BI values of preys are measured with quantitative protein analytical methods, such as WB or MS ( Fig. 1A ) ( 19 , 20 )⁠. While the qualitative proof of concept of nHU was demonstrated before, here, we develop it into a quantitative assay ( 19 )⁠.…”

Section: Resultsmentioning

confidence: 99%

Native holdup (nHU) to measure binding affinities from cell extracts

et al. 2022

View full text Add to dashboard Cite

Characterizing macromolecular interactions is essential for understanding cellular processes, yet most methods currently used to detect protein interactions from cells are qualitative. Here, we introduce the native holdup (nHU) approach to estimate equilibrium binding constants of protein interactions directly from cell extracts. Compared to other pull-down–based assays, nHU requires less sample preparation and can be coupled to any analytical methods as readouts, such as Western blotting or mass spectrometry. We use nHU to explore interactions of SNX27, a cargo adaptor of the retromer complex and find good agreement between in vitro affinities and those measured directly from cell extracts using nHU. We discuss the strengths and limitations of nHU and provide simple protocols that can be implemented in most laboratories.

show abstract

Promiscuity mapping of the S100 protein family using a high-throughput holdup assay

Cited by 10 publications

References 35 publications

Calcium-Bound S100P Protein Is a Promiscuous Binding Partner of the Four-Helical Cytokines

Calcium-Bound S100P Protein Is a Promiscuous Binding Partner of the Four-Helical Cytokines

Calcium-Bound S100P Protein is a Promiscuous Binding Partner of the Four-Helical Cytokines

Native holdup (nHU) to measure binding affinities from cell extracts

Contact Info

Product

Resources

About