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pH-Dependent Interaction between C-Peptide and Phospholipid Bicelles.

Unnerståle S, Mäler L - J Biophys (2012)

Bottom Line: The results demonstrate that C-peptide is largely unstructured independent of pH, but that a weak structural induction towards a short stretch of β-sheet is induced at low pH, corresponding to the isoelectric point of the peptide.C-peptide does not undergo a large structural rearrangement as a consequence of lipid interaction, which indicates that the folding and binding are uncoupled.In vivo, local variations in environment, including pH, may cause C-peptide to associate with lipids, which may affect the aggregation state of the peptide.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Biophysics, Center for Biomembrane Research, The Arrhenius Laboratories for Natural Sciences, Stockholm University, 106 91 Stockholm, Sweden.

ABSTRACT
C-peptide is the connecting peptide between the A and B chains of insulin in proinsulin. In this paper, we investigate the interaction between C-peptide and phospholipid bicelles, by circular dichroism and nuclear magnetic resonance spectroscopy, and in particular the pH dependence of this interaction. The results demonstrate that C-peptide is largely unstructured independent of pH, but that a weak structural induction towards a short stretch of β-sheet is induced at low pH, corresponding to the isoelectric point of the peptide. Furthermore, it is demonstrated that C-peptide associates with neutral phospholipid bicelles as well as acidic phospholipid bicelles at this low pH. C-peptide does not undergo a large structural rearrangement as a consequence of lipid interaction, which indicates that the folding and binding are uncoupled. In vivo, local variations in environment, including pH, may cause C-peptide to associate with lipids, which may affect the aggregation state of the peptide.

No MeSH data available.


Related in: MedlinePlus

When lowering the pH to 3.2 (left panel), small structural rearrangements are induced that facilitate interaction with phospholipid bicelles. The interaction with bicelles further influences the structure. At pH 5.8, on the other hand, (right panel), no structural arrangements are seen upon addition of bicelles and the interaction is much weaker.
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fig6: When lowering the pH to 3.2 (left panel), small structural rearrangements are induced that facilitate interaction with phospholipid bicelles. The interaction with bicelles further influences the structure. At pH 5.8, on the other hand, (right panel), no structural arrangements are seen upon addition of bicelles and the interaction is much weaker.

Mentions: In this study, we wished to elucidate the effect of pH on C-peptide structure and lipid interactions. Previous studies have examined the effects of lipids on C-peptide structure at pH 5 or higher, and neither CD spectroscopy or size exclusion chromatography has revealed any lipid interaction [12]. Hence, it was then concluded that stable conformation-dependent interactions of C-peptide with lipid membranes are unlikely to occur. Biological effects of C-peptide, protecting against diabetic complications, are mediated by interaction with insulin or interaction with membrane via specific and/or nonspecific membrane interaction. Most studies support specific interactions with a, yet to be found, GPCR [40, 41]. However, the D-enantiomer of C-peptide has the same biological activity as the L-enantiomer [42], which suggests that other receptor-independent interactions are important for function. Formation of cation-selective channels in lipid bilayers [43] also suggests a more nonspecific interaction. Hence, we find it valuable to investigate nonspecific interactions between C-peptide and the membrane as a part of C-peptides protective function. We have previously demonstrated that, at low pH, C-peptide has the ability to form β-sheet-like aggregates at low detergent concentrations and α-helical structure in SDS micelles [15], indicating that pH is important for structural induction. Thus, the structure and lipid interaction of C-peptide was in the present study also examined at a lower pH close to the pI of the C-peptide. From the results, we see that C-peptide favors a lipid interaction at low pH, when the peptide is neutral, (around 65% of the peptide is associated with bicelles at pH 3.2), suggesting that the relationship between electrostatic and hydrophobic interactions is important for this process (Figure 6). By decreasing the pH, small structural rearrangements in predominately the N-terminal and in the amino acid stretch between V10 and L12 are induced, that facilitate lipid interaction. Upon addition of bicelles, these structural preferences are stabilized. The structural rearrangements of C-peptide, as judged from both CD and NMR spectroscopy are not large, and thus, C-peptide represents a group of membrane interacting peptides that do not appear to undergo large structural changes upon membrane binding. This behavior has previously been observed for, for example, the interaction between the opioid receptor peptide ligands (dynorphins) and bicelles, which did not cause any structural induction in the peptide ligands [34]. It appears that lack of structure induction is not a conclusive way of demonstrating lack of peptide-membrane interaction. Sometimes local and transient structural preferences in an ensemble of peptides dictate function [44]. This is similar to the recent findings that even protein-protein interactions may not always lead to well-formed secondary or tertiary structures, but indicates a novel mode of action of these intrinsically disordered proteins [45, 46].


pH-Dependent Interaction between C-Peptide and Phospholipid Bicelles.

Unnerståle S, Mäler L - J Biophys (2012)

When lowering the pH to 3.2 (left panel), small structural rearrangements are induced that facilitate interaction with phospholipid bicelles. The interaction with bicelles further influences the structure. At pH 5.8, on the other hand, (right panel), no structural arrangements are seen upon addition of bicelles and the interaction is much weaker.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3405660&req=5

fig6: When lowering the pH to 3.2 (left panel), small structural rearrangements are induced that facilitate interaction with phospholipid bicelles. The interaction with bicelles further influences the structure. At pH 5.8, on the other hand, (right panel), no structural arrangements are seen upon addition of bicelles and the interaction is much weaker.
Mentions: In this study, we wished to elucidate the effect of pH on C-peptide structure and lipid interactions. Previous studies have examined the effects of lipids on C-peptide structure at pH 5 or higher, and neither CD spectroscopy or size exclusion chromatography has revealed any lipid interaction [12]. Hence, it was then concluded that stable conformation-dependent interactions of C-peptide with lipid membranes are unlikely to occur. Biological effects of C-peptide, protecting against diabetic complications, are mediated by interaction with insulin or interaction with membrane via specific and/or nonspecific membrane interaction. Most studies support specific interactions with a, yet to be found, GPCR [40, 41]. However, the D-enantiomer of C-peptide has the same biological activity as the L-enantiomer [42], which suggests that other receptor-independent interactions are important for function. Formation of cation-selective channels in lipid bilayers [43] also suggests a more nonspecific interaction. Hence, we find it valuable to investigate nonspecific interactions between C-peptide and the membrane as a part of C-peptides protective function. We have previously demonstrated that, at low pH, C-peptide has the ability to form β-sheet-like aggregates at low detergent concentrations and α-helical structure in SDS micelles [15], indicating that pH is important for structural induction. Thus, the structure and lipid interaction of C-peptide was in the present study also examined at a lower pH close to the pI of the C-peptide. From the results, we see that C-peptide favors a lipid interaction at low pH, when the peptide is neutral, (around 65% of the peptide is associated with bicelles at pH 3.2), suggesting that the relationship between electrostatic and hydrophobic interactions is important for this process (Figure 6). By decreasing the pH, small structural rearrangements in predominately the N-terminal and in the amino acid stretch between V10 and L12 are induced, that facilitate lipid interaction. Upon addition of bicelles, these structural preferences are stabilized. The structural rearrangements of C-peptide, as judged from both CD and NMR spectroscopy are not large, and thus, C-peptide represents a group of membrane interacting peptides that do not appear to undergo large structural changes upon membrane binding. This behavior has previously been observed for, for example, the interaction between the opioid receptor peptide ligands (dynorphins) and bicelles, which did not cause any structural induction in the peptide ligands [34]. It appears that lack of structure induction is not a conclusive way of demonstrating lack of peptide-membrane interaction. Sometimes local and transient structural preferences in an ensemble of peptides dictate function [44]. This is similar to the recent findings that even protein-protein interactions may not always lead to well-formed secondary or tertiary structures, but indicates a novel mode of action of these intrinsically disordered proteins [45, 46].

Bottom Line: The results demonstrate that C-peptide is largely unstructured independent of pH, but that a weak structural induction towards a short stretch of β-sheet is induced at low pH, corresponding to the isoelectric point of the peptide.C-peptide does not undergo a large structural rearrangement as a consequence of lipid interaction, which indicates that the folding and binding are uncoupled.In vivo, local variations in environment, including pH, may cause C-peptide to associate with lipids, which may affect the aggregation state of the peptide.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Biophysics, Center for Biomembrane Research, The Arrhenius Laboratories for Natural Sciences, Stockholm University, 106 91 Stockholm, Sweden.

ABSTRACT
C-peptide is the connecting peptide between the A and B chains of insulin in proinsulin. In this paper, we investigate the interaction between C-peptide and phospholipid bicelles, by circular dichroism and nuclear magnetic resonance spectroscopy, and in particular the pH dependence of this interaction. The results demonstrate that C-peptide is largely unstructured independent of pH, but that a weak structural induction towards a short stretch of β-sheet is induced at low pH, corresponding to the isoelectric point of the peptide. Furthermore, it is demonstrated that C-peptide associates with neutral phospholipid bicelles as well as acidic phospholipid bicelles at this low pH. C-peptide does not undergo a large structural rearrangement as a consequence of lipid interaction, which indicates that the folding and binding are uncoupled. In vivo, local variations in environment, including pH, may cause C-peptide to associate with lipids, which may affect the aggregation state of the peptide.

No MeSH data available.


Related in: MedlinePlus