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Charge environments around phosphorylation sites in proteins.

Kitchen J, Saunders RE, Warwicker J - BMC Struct. Biol. (2008)

Bottom Line: A significant fraction of phosphorylated sites appear to be electrostatically stabilised, largely through interaction with sidechains.In this study, an easy to implement procedure is developed that could contribute to the identification of phospho-acceptor sites associated with charge-charge interactions and conformational change.Our analysis of the Phospho.ELM dataset also shows evidence for mediation of phosphorylation effects through (i) conformational change associated with making a solvent inaccessible phospho-acceptor site accessible, and (ii) modulation of protein-protein interactions.

View Article: PubMed Central - HTML - PubMed

Affiliation: Faculty of Life Sciences, University of Manchester, Michael Smith Building, Oxford Road, Manchester M13 9PT, UK. j.kitchen@student.manchester.ac.uk

ABSTRACT

Background: Phosphorylation is a central feature in many biological processes. Structural analyses have identified the importance of charge-charge interactions, for example mediating phosphorylation-driven allosteric change and protein binding to phosphopeptides. Here, we examine computationally the prevalence of charge stabilisation around phosphorylated sites in the structural database, through comparison with locations that are not phosphorylated in the same structures.

Results: A significant fraction of phosphorylated sites appear to be electrostatically stabilised, largely through interaction with sidechains. Some examples of stabilisation across a subunit interface are evident from calculations with biological units. When considering the immediately surrounding environment, in many cases favourable interactions are only apparent after conformational change that accompanies phosphorylation. A simple calculation of potential interactions at longer-range, applied to non-phosphorylated structures, recovers the separation exhibited by phosphorylated structures. In a study of sites in the Phospho.ELM dataset, for which structural annotation is provided by non-phosphorylated proteins, there is little separation of the known phospho-acceptor sites relative to background, even using the wider interaction radius. However, there are differences in the distributions of patch polarity for acceptor and background sites in the Phospho.ELM dataset.

Conclusion: In this study, an easy to implement procedure is developed that could contribute to the identification of phospho-acceptor sites associated with charge-charge interactions and conformational change. Since the method gives information about potential anchoring interactions subsequent to phosphorylation, it could be combined with simulations that probe conformational change. Our analysis of the Phospho.ELM dataset also shows evidence for mediation of phosphorylation effects through (i) conformational change associated with making a solvent inaccessible phospho-acceptor site accessible, and (ii) modulation of protein-protein interactions.

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Searching for favourable interactions around phospho-acceptor sites in proteins that are not phosphorylated. Calculations were made with the non-phosphorylated set of proteins, from the phosphorylated/non-phosphorylated structure pairs, many of which undergo conformational change upon phosphorylation. A search was made, within a given radius, for the most positive potential peak around each site (real/phos or background/non-phos). Interaction energy is given for a single unit negative charge in the positive potential field. (a) Radius = 5 Å. (b) Radius = 10 Å. (c) Radius = 30 Å.
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Figure 8: Searching for favourable interactions around phospho-acceptor sites in proteins that are not phosphorylated. Calculations were made with the non-phosphorylated set of proteins, from the phosphorylated/non-phosphorylated structure pairs, many of which undergo conformational change upon phosphorylation. A search was made, within a given radius, for the most positive potential peak around each site (real/phos or background/non-phos). Interaction energy is given for a single unit negative charge in the positive potential field. (a) Radius = 5 Å. (b) Radius = 10 Å. (c) Radius = 30 Å.

Mentions: Since mainchain conformational change plays a role in some phosphorylations, we adapted previous work that studied electrostatic potential values on a solvent accessible shell [55]. In this case we are looking for the most favourable interacting site (highest positive potential) within a given radius of each site (centred on Ser OG, Thr OG1 or Tyr OH). Figure 8 shows this analysis for the non-phosphorylated coordinate sets of the previous section, at three radii. At 30 Å radius, the interaction values are generally favourable and there is little separation between the real and background sets, since the large search radius links all Ser, Thr, Tyr sites to favourable patches for charge interaction. Of more interest are the smaller radii. Some separation is apparent at 5 Å radius, which increases at 10 Å radius, and is largely associated with the proteins (including kinases) that underpin separation in earlier Figures (e.g. Figure 7a). The procedure of electrostatic peak finding within a sphere around a phospho-acceptor site, but for non-phosphorylated molecules, largely recovers the result of calculations with phosphorylated coordinate sets. This occurs without knowledge of the loops that undergo conformational change. The method could therefore aid identification of such regions, and allied with simulation methods [34], provide structural models for conformational change that is coupled to phosphorylation.


Charge environments around phosphorylation sites in proteins.

Kitchen J, Saunders RE, Warwicker J - BMC Struct. Biol. (2008)

Searching for favourable interactions around phospho-acceptor sites in proteins that are not phosphorylated. Calculations were made with the non-phosphorylated set of proteins, from the phosphorylated/non-phosphorylated structure pairs, many of which undergo conformational change upon phosphorylation. A search was made, within a given radius, for the most positive potential peak around each site (real/phos or background/non-phos). Interaction energy is given for a single unit negative charge in the positive potential field. (a) Radius = 5 Å. (b) Radius = 10 Å. (c) Radius = 30 Å.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2291461&req=5

Figure 8: Searching for favourable interactions around phospho-acceptor sites in proteins that are not phosphorylated. Calculations were made with the non-phosphorylated set of proteins, from the phosphorylated/non-phosphorylated structure pairs, many of which undergo conformational change upon phosphorylation. A search was made, within a given radius, for the most positive potential peak around each site (real/phos or background/non-phos). Interaction energy is given for a single unit negative charge in the positive potential field. (a) Radius = 5 Å. (b) Radius = 10 Å. (c) Radius = 30 Å.
Mentions: Since mainchain conformational change plays a role in some phosphorylations, we adapted previous work that studied electrostatic potential values on a solvent accessible shell [55]. In this case we are looking for the most favourable interacting site (highest positive potential) within a given radius of each site (centred on Ser OG, Thr OG1 or Tyr OH). Figure 8 shows this analysis for the non-phosphorylated coordinate sets of the previous section, at three radii. At 30 Å radius, the interaction values are generally favourable and there is little separation between the real and background sets, since the large search radius links all Ser, Thr, Tyr sites to favourable patches for charge interaction. Of more interest are the smaller radii. Some separation is apparent at 5 Å radius, which increases at 10 Å radius, and is largely associated with the proteins (including kinases) that underpin separation in earlier Figures (e.g. Figure 7a). The procedure of electrostatic peak finding within a sphere around a phospho-acceptor site, but for non-phosphorylated molecules, largely recovers the result of calculations with phosphorylated coordinate sets. This occurs without knowledge of the loops that undergo conformational change. The method could therefore aid identification of such regions, and allied with simulation methods [34], provide structural models for conformational change that is coupled to phosphorylation.

Bottom Line: A significant fraction of phosphorylated sites appear to be electrostatically stabilised, largely through interaction with sidechains.In this study, an easy to implement procedure is developed that could contribute to the identification of phospho-acceptor sites associated with charge-charge interactions and conformational change.Our analysis of the Phospho.ELM dataset also shows evidence for mediation of phosphorylation effects through (i) conformational change associated with making a solvent inaccessible phospho-acceptor site accessible, and (ii) modulation of protein-protein interactions.

View Article: PubMed Central - HTML - PubMed

Affiliation: Faculty of Life Sciences, University of Manchester, Michael Smith Building, Oxford Road, Manchester M13 9PT, UK. j.kitchen@student.manchester.ac.uk

ABSTRACT

Background: Phosphorylation is a central feature in many biological processes. Structural analyses have identified the importance of charge-charge interactions, for example mediating phosphorylation-driven allosteric change and protein binding to phosphopeptides. Here, we examine computationally the prevalence of charge stabilisation around phosphorylated sites in the structural database, through comparison with locations that are not phosphorylated in the same structures.

Results: A significant fraction of phosphorylated sites appear to be electrostatically stabilised, largely through interaction with sidechains. Some examples of stabilisation across a subunit interface are evident from calculations with biological units. When considering the immediately surrounding environment, in many cases favourable interactions are only apparent after conformational change that accompanies phosphorylation. A simple calculation of potential interactions at longer-range, applied to non-phosphorylated structures, recovers the separation exhibited by phosphorylated structures. In a study of sites in the Phospho.ELM dataset, for which structural annotation is provided by non-phosphorylated proteins, there is little separation of the known phospho-acceptor sites relative to background, even using the wider interaction radius. However, there are differences in the distributions of patch polarity for acceptor and background sites in the Phospho.ELM dataset.

Conclusion: In this study, an easy to implement procedure is developed that could contribute to the identification of phospho-acceptor sites associated with charge-charge interactions and conformational change. Since the method gives information about potential anchoring interactions subsequent to phosphorylation, it could be combined with simulations that probe conformational change. Our analysis of the Phospho.ELM dataset also shows evidence for mediation of phosphorylation effects through (i) conformational change associated with making a solvent inaccessible phospho-acceptor site accessible, and (ii) modulation of protein-protein interactions.

Show MeSH
Related in: MedlinePlus