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Tandem Domains with Tuned Interactions Are a Powerful Biological Design Principle.

Nussinov R, Tsai CJ - PLoS Biol. (2015)

Bottom Line: Allosteric effects of mutations, ligand binding, or post-translational modifications on protein function occur through changes to the protein's shape, or conformation.In a new PLOS Biology paper, Melacini and colleagues describe a novel model of protein regulation, the "Double-Conformational Selection Model", which demonstrates how two tandem ligand-binding domains interact to regulate protein function.Here we explain how tandem domains with tuned interactions-but not single domains-can provide a blueprint for sensitive activation sensors within a narrow window of ligand concentration, thereby promoting signaling control.

View Article: PubMed Central - PubMed

Affiliation: Cancer and Inflammation Program, Leidos Biomedical Research, Inc., Frederick, Maryland, United States of America.

ABSTRACT
Allosteric effects of mutations, ligand binding, or post-translational modifications on protein function occur through changes to the protein's shape, or conformation. In a cell, there are many copies of the same protein, all experiencing these perturbations in a dynamic fashion and fluctuating through different conformations and activity states. According to the "conformational selection and population shift" theory, ligand binding selects a particular conformation. This perturbs the ensemble and induces a population shift. In a new PLOS Biology paper, Melacini and colleagues describe a novel model of protein regulation, the "Double-Conformational Selection Model", which demonstrates how two tandem ligand-binding domains interact to regulate protein function. Here we explain how tandem domains with tuned interactions-but not single domains-can provide a blueprint for sensitive activation sensors within a narrow window of ligand concentration, thereby promoting signaling control.

Show MeSH

Related in: MedlinePlus

The role of conformational selection and population shift in regulation of PKA activity.(A) We start with four independent population states of PKA regulatory subunit (RIα) with two tandem cAMP binding domains (CBD-A and CBD-B) either in active (small red oval) or inactive (small green oval) states, presumably in open arrangements that have been verified by the NMR (nuclear magnetic resonance) experiments [5]. The top panel shows how the catalytic subunit of PKA (large orange oval) is secured in the inhibited (OFF, inactive) state by the regulatory subunit in the absence of cAMP. First, through conformation selection, the kinase domain only binds to inactive CBDs. The two tandem CBDs can exist in three possible populations (inactive–inactive, active–inactive, and inactive–active), with different binding affinities between them. Then, thanks to the tandem arrangement of the CBDs and/or via the allosteric influence resulting from the inactive CBD binding, the other, active CBD states are further stabilized in the inactive form when bound to the kinase subunit as indicated by the blue curved arrows. To ensure limited activity in the absence of cAMP, the formation of a tetramer by dimerization (indicated by the arrows) further effectively reduces the free active kinase subunit. The bottom panel shows how cAMP (tiny cyan oval) maximizes the concentration of free catalytic kinase through its binding to the PKA regulatory subunit. First, the panel shows that cAMP selectively binds to inactive CBDs only. Then, the catalytic subunit is only permitted to bind to the population of the regulatory subunit with both inactive CBDs. Next, the cAMP-bound active CBD allosterically shifts the other, inactive CBD into active conformation (indicated by the arrows), which allows cAMP to occupy both binding sites in CBD. (B) Large (inter-CBDs) and small (intra-CBD) conformational changes can be seen through the superposition of CBD-A domains taken from the open tandem CBDs (green) bound to the catalytic kinase subunit (yellow) (PDB: 1 rgs) and the closed form of the regulatory subunit (red) (PDB: 2 qcs). The steric collision between the catalytic kinase subunit and the closed CBD-B domain seen in the figure reveals why the large conformational change further destabilizes substantially the binding affinity between CBD-A and the catalytic subunit, which has already been reduced by the small intra-CBD conformational change due to cAMP binding.
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pbio.1002306.g001: The role of conformational selection and population shift in regulation of PKA activity.(A) We start with four independent population states of PKA regulatory subunit (RIα) with two tandem cAMP binding domains (CBD-A and CBD-B) either in active (small red oval) or inactive (small green oval) states, presumably in open arrangements that have been verified by the NMR (nuclear magnetic resonance) experiments [5]. The top panel shows how the catalytic subunit of PKA (large orange oval) is secured in the inhibited (OFF, inactive) state by the regulatory subunit in the absence of cAMP. First, through conformation selection, the kinase domain only binds to inactive CBDs. The two tandem CBDs can exist in three possible populations (inactive–inactive, active–inactive, and inactive–active), with different binding affinities between them. Then, thanks to the tandem arrangement of the CBDs and/or via the allosteric influence resulting from the inactive CBD binding, the other, active CBD states are further stabilized in the inactive form when bound to the kinase subunit as indicated by the blue curved arrows. To ensure limited activity in the absence of cAMP, the formation of a tetramer by dimerization (indicated by the arrows) further effectively reduces the free active kinase subunit. The bottom panel shows how cAMP (tiny cyan oval) maximizes the concentration of free catalytic kinase through its binding to the PKA regulatory subunit. First, the panel shows that cAMP selectively binds to inactive CBDs only. Then, the catalytic subunit is only permitted to bind to the population of the regulatory subunit with both inactive CBDs. Next, the cAMP-bound active CBD allosterically shifts the other, inactive CBD into active conformation (indicated by the arrows), which allows cAMP to occupy both binding sites in CBD. (B) Large (inter-CBDs) and small (intra-CBD) conformational changes can be seen through the superposition of CBD-A domains taken from the open tandem CBDs (green) bound to the catalytic kinase subunit (yellow) (PDB: 1 rgs) and the closed form of the regulatory subunit (red) (PDB: 2 qcs). The steric collision between the catalytic kinase subunit and the closed CBD-B domain seen in the figure reveals why the large conformational change further destabilizes substantially the binding affinity between CBD-A and the catalytic subunit, which has already been reduced by the small intra-CBD conformational change due to cAMP binding.

Mentions: How can conformational selection lead to a steep activation of a protein within a narrow range of ligand concentration? The regulation of protein kinase A (PKA) by its regulatory subunit (RIα) suggests that evolution has come up with an elegant solution: the engineered RIα homologous tandem cAMP binding domains (CBD-A, CBD-B) organization. When not bound to cAMP, RIα is in an open conformation, with weak interactions between the domains. With higher affinity, CBD-B is the first to bind cAMP. As Melacini and his colleagues show [5], binding shifts the RIα ensemble. This has two consequences: it increases the population of a cAMP-compatible conformation in CBD-A, which now also binds cAMP, and in particular strengthens the weak CBD-A/CBD-B domain–domain interactions. Even though the cAMP-elicited conformational change in each of the domains is minor, the global conformational change of RIα from an open to a closed conformation is large. This is important since the stable closed RIα conformation has steric conflicts with PKA. The more stable closed conformation now shifts the equilibrium away from the open state, and the freed PKA is activated. Note that even in the absence of cAMP, holo-like closed RIα conformations are present in the ensemble; however, the populations of these states are functionally insignificant. Why is such a mechanism advantageous to the cell? It allows PKA an efficient cAMP concentration-sensitive molecular response. Protein design efforts aiming at efficient activation through sharp concentration transitions may consider exploiting such a strategy. Fig 1 illustrates how this mechanism works.


Tandem Domains with Tuned Interactions Are a Powerful Biological Design Principle.

Nussinov R, Tsai CJ - PLoS Biol. (2015)

The role of conformational selection and population shift in regulation of PKA activity.(A) We start with four independent population states of PKA regulatory subunit (RIα) with two tandem cAMP binding domains (CBD-A and CBD-B) either in active (small red oval) or inactive (small green oval) states, presumably in open arrangements that have been verified by the NMR (nuclear magnetic resonance) experiments [5]. The top panel shows how the catalytic subunit of PKA (large orange oval) is secured in the inhibited (OFF, inactive) state by the regulatory subunit in the absence of cAMP. First, through conformation selection, the kinase domain only binds to inactive CBDs. The two tandem CBDs can exist in three possible populations (inactive–inactive, active–inactive, and inactive–active), with different binding affinities between them. Then, thanks to the tandem arrangement of the CBDs and/or via the allosteric influence resulting from the inactive CBD binding, the other, active CBD states are further stabilized in the inactive form when bound to the kinase subunit as indicated by the blue curved arrows. To ensure limited activity in the absence of cAMP, the formation of a tetramer by dimerization (indicated by the arrows) further effectively reduces the free active kinase subunit. The bottom panel shows how cAMP (tiny cyan oval) maximizes the concentration of free catalytic kinase through its binding to the PKA regulatory subunit. First, the panel shows that cAMP selectively binds to inactive CBDs only. Then, the catalytic subunit is only permitted to bind to the population of the regulatory subunit with both inactive CBDs. Next, the cAMP-bound active CBD allosterically shifts the other, inactive CBD into active conformation (indicated by the arrows), which allows cAMP to occupy both binding sites in CBD. (B) Large (inter-CBDs) and small (intra-CBD) conformational changes can be seen through the superposition of CBD-A domains taken from the open tandem CBDs (green) bound to the catalytic kinase subunit (yellow) (PDB: 1 rgs) and the closed form of the regulatory subunit (red) (PDB: 2 qcs). The steric collision between the catalytic kinase subunit and the closed CBD-B domain seen in the figure reveals why the large conformational change further destabilizes substantially the binding affinity between CBD-A and the catalytic subunit, which has already been reduced by the small intra-CBD conformational change due to cAMP binding.
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pbio.1002306.g001: The role of conformational selection and population shift in regulation of PKA activity.(A) We start with four independent population states of PKA regulatory subunit (RIα) with two tandem cAMP binding domains (CBD-A and CBD-B) either in active (small red oval) or inactive (small green oval) states, presumably in open arrangements that have been verified by the NMR (nuclear magnetic resonance) experiments [5]. The top panel shows how the catalytic subunit of PKA (large orange oval) is secured in the inhibited (OFF, inactive) state by the regulatory subunit in the absence of cAMP. First, through conformation selection, the kinase domain only binds to inactive CBDs. The two tandem CBDs can exist in three possible populations (inactive–inactive, active–inactive, and inactive–active), with different binding affinities between them. Then, thanks to the tandem arrangement of the CBDs and/or via the allosteric influence resulting from the inactive CBD binding, the other, active CBD states are further stabilized in the inactive form when bound to the kinase subunit as indicated by the blue curved arrows. To ensure limited activity in the absence of cAMP, the formation of a tetramer by dimerization (indicated by the arrows) further effectively reduces the free active kinase subunit. The bottom panel shows how cAMP (tiny cyan oval) maximizes the concentration of free catalytic kinase through its binding to the PKA regulatory subunit. First, the panel shows that cAMP selectively binds to inactive CBDs only. Then, the catalytic subunit is only permitted to bind to the population of the regulatory subunit with both inactive CBDs. Next, the cAMP-bound active CBD allosterically shifts the other, inactive CBD into active conformation (indicated by the arrows), which allows cAMP to occupy both binding sites in CBD. (B) Large (inter-CBDs) and small (intra-CBD) conformational changes can be seen through the superposition of CBD-A domains taken from the open tandem CBDs (green) bound to the catalytic kinase subunit (yellow) (PDB: 1 rgs) and the closed form of the regulatory subunit (red) (PDB: 2 qcs). The steric collision between the catalytic kinase subunit and the closed CBD-B domain seen in the figure reveals why the large conformational change further destabilizes substantially the binding affinity between CBD-A and the catalytic subunit, which has already been reduced by the small intra-CBD conformational change due to cAMP binding.
Mentions: How can conformational selection lead to a steep activation of a protein within a narrow range of ligand concentration? The regulation of protein kinase A (PKA) by its regulatory subunit (RIα) suggests that evolution has come up with an elegant solution: the engineered RIα homologous tandem cAMP binding domains (CBD-A, CBD-B) organization. When not bound to cAMP, RIα is in an open conformation, with weak interactions between the domains. With higher affinity, CBD-B is the first to bind cAMP. As Melacini and his colleagues show [5], binding shifts the RIα ensemble. This has two consequences: it increases the population of a cAMP-compatible conformation in CBD-A, which now also binds cAMP, and in particular strengthens the weak CBD-A/CBD-B domain–domain interactions. Even though the cAMP-elicited conformational change in each of the domains is minor, the global conformational change of RIα from an open to a closed conformation is large. This is important since the stable closed RIα conformation has steric conflicts with PKA. The more stable closed conformation now shifts the equilibrium away from the open state, and the freed PKA is activated. Note that even in the absence of cAMP, holo-like closed RIα conformations are present in the ensemble; however, the populations of these states are functionally insignificant. Why is such a mechanism advantageous to the cell? It allows PKA an efficient cAMP concentration-sensitive molecular response. Protein design efforts aiming at efficient activation through sharp concentration transitions may consider exploiting such a strategy. Fig 1 illustrates how this mechanism works.

Bottom Line: Allosteric effects of mutations, ligand binding, or post-translational modifications on protein function occur through changes to the protein's shape, or conformation.In a new PLOS Biology paper, Melacini and colleagues describe a novel model of protein regulation, the "Double-Conformational Selection Model", which demonstrates how two tandem ligand-binding domains interact to regulate protein function.Here we explain how tandem domains with tuned interactions-but not single domains-can provide a blueprint for sensitive activation sensors within a narrow window of ligand concentration, thereby promoting signaling control.

View Article: PubMed Central - PubMed

Affiliation: Cancer and Inflammation Program, Leidos Biomedical Research, Inc., Frederick, Maryland, United States of America.

ABSTRACT
Allosteric effects of mutations, ligand binding, or post-translational modifications on protein function occur through changes to the protein's shape, or conformation. In a cell, there are many copies of the same protein, all experiencing these perturbations in a dynamic fashion and fluctuating through different conformations and activity states. According to the "conformational selection and population shift" theory, ligand binding selects a particular conformation. This perturbs the ensemble and induces a population shift. In a new PLOS Biology paper, Melacini and colleagues describe a novel model of protein regulation, the "Double-Conformational Selection Model", which demonstrates how two tandem ligand-binding domains interact to regulate protein function. Here we explain how tandem domains with tuned interactions-but not single domains-can provide a blueprint for sensitive activation sensors within a narrow window of ligand concentration, thereby promoting signaling control.

Show MeSH
Related in: MedlinePlus