Limits...
Headgroup-dependent membrane catalysis of apelin-receptor interactions is likely.

Langelaan DN, Rainey JK - J Phys Chem B (2009)

Bottom Line: Type I beta-turns are initiated between R6 and L9, and a well-defined type IV beta-turn is initiated at S10.Furthermore, binding of apelin-17 to SDS micelles causes structuring of M15-F17, with no evidence for direct binding of this region to the micelles.These results are placed into the context of the membrane catalysis hypothesis for peptide-receptor binding, and a hypothetical mechanism of APJ binding and activation by apelin is advanced.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, Nova Scotia B3H 1X5, Canada.

ABSTRACT
Apelin is the peptidic ligand for the G-protein-coupled receptor APJ. The apelin-APJ system is important in cardiovascular regulation, fluid homeostasis, and angiogenesis, among other roles. In this study, we investigate interactions between apelin and membrane-mimetic micelles of the detergents sodium dodecyl sulfate (SDS), dodecylphosphocholine (DPC), and 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-rac-(1-glycerol)] (LPPG). Far-ultraviolet circular dichroism spectropolarimetry and diffusion-ordered spectroscopy indicate that apelin peptides bind to micelles of the anionic detergents SDS and LPPG much more favorably than to zwitterionic DPC micelles. Nuclear magnetic resonance spectroscopy allowed full characterization of the interactions of apelin-17 with SDS micelles. Titration with paramagnetic agents and structural determination of apelin-17 in SDS indicate that R6-K12 is highly structured, with R6-L9 directly interacting with headgroups of the micelle. Type I beta-turns are initiated between R6 and L9, and a well-defined type IV beta-turn is initiated at S10. Furthermore, binding of apelin-17 to SDS micelles causes structuring of M15-F17, with no evidence for direct binding of this region to the micelles. These results are placed into the context of the membrane catalysis hypothesis for peptide-receptor binding, and a hypothetical mechanism of APJ binding and activation by apelin is advanced.

Show MeSH
Apelin-17 secondary chemical shifts (Δδ = δ(obsd) − δ(random coil)(29)) for Hα, Cα, Hβ, and Cβ (the prime symbol denotes a degenerate shift) at 35 °C in buffer(15) (20 mM CD3COO−, pH 5.00 ± 0.05), SDS micelles, and DPC micelles. Horizontal lines show the Δδ cutoff significant for secondary structuring(31) (note that Cα and Cβ for Pro have identified significance ranges of ±4 ppm).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC2736645&req=5

fig2: Apelin-17 secondary chemical shifts (Δδ = δ(obsd) − δ(random coil)(29)) for Hα, Cα, Hβ, and Cβ (the prime symbol denotes a degenerate shift) at 35 °C in buffer(15) (20 mM CD3COO−, pH 5.00 ± 0.05), SDS micelles, and DPC micelles. Horizontal lines show the Δδ cutoff significant for secondary structuring(31) (note that Cα and Cβ for Pro have identified significance ranges of ±4 ppm).

Mentions: Nearly complete assignment of 1H, 13C, and 15N was obtained for apelin-17 (Table S1 in the Supporting Information) in the presence of both SDS and DPC micelles using standard methods,(53) allowing comparison of Δδ32 for Hα, Cα, Hβ, and Cβ nuclei in the presence of SDS and DPC micelles to those in buffer that we previously reported(15) (Figure 2). For both Hα and Cβ, there is no clear relationship between the value of the secondary chemical shift and the condition used. However, these Δδ values are significantly different in the presence of SDS micelles compared to those in either the buffer or DPC conditions over K1−K12, while Δδ values in the C-terminal of apelin-17 show little perturbation in the presence of micelles. The large changes in Δδ values of Cα and Hβ atoms in the presence of SDS suggest that apelin-17 is binding to the SDS micelles rather than forming a different, canonical secondary structure since Hα and Cβ would also be expected to have a consistent Δδ perturbation with secondary structure formation. Determination of the effect of LPPG micelles on Δδ values was not possible as deuterated LPPG is not available, making spectral assignment infeasible for apelin without isotopic enrichment.


Headgroup-dependent membrane catalysis of apelin-receptor interactions is likely.

Langelaan DN, Rainey JK - J Phys Chem B (2009)

Apelin-17 secondary chemical shifts (Δδ = δ(obsd) − δ(random coil)(29)) for Hα, Cα, Hβ, and Cβ (the prime symbol denotes a degenerate shift) at 35 °C in buffer(15) (20 mM CD3COO−, pH 5.00 ± 0.05), SDS micelles, and DPC micelles. Horizontal lines show the Δδ cutoff significant for secondary structuring(31) (note that Cα and Cβ for Pro have identified significance ranges of ±4 ppm).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2736645&req=5

fig2: Apelin-17 secondary chemical shifts (Δδ = δ(obsd) − δ(random coil)(29)) for Hα, Cα, Hβ, and Cβ (the prime symbol denotes a degenerate shift) at 35 °C in buffer(15) (20 mM CD3COO−, pH 5.00 ± 0.05), SDS micelles, and DPC micelles. Horizontal lines show the Δδ cutoff significant for secondary structuring(31) (note that Cα and Cβ for Pro have identified significance ranges of ±4 ppm).
Mentions: Nearly complete assignment of 1H, 13C, and 15N was obtained for apelin-17 (Table S1 in the Supporting Information) in the presence of both SDS and DPC micelles using standard methods,(53) allowing comparison of Δδ32 for Hα, Cα, Hβ, and Cβ nuclei in the presence of SDS and DPC micelles to those in buffer that we previously reported(15) (Figure 2). For both Hα and Cβ, there is no clear relationship between the value of the secondary chemical shift and the condition used. However, these Δδ values are significantly different in the presence of SDS micelles compared to those in either the buffer or DPC conditions over K1−K12, while Δδ values in the C-terminal of apelin-17 show little perturbation in the presence of micelles. The large changes in Δδ values of Cα and Hβ atoms in the presence of SDS suggest that apelin-17 is binding to the SDS micelles rather than forming a different, canonical secondary structure since Hα and Cβ would also be expected to have a consistent Δδ perturbation with secondary structure formation. Determination of the effect of LPPG micelles on Δδ values was not possible as deuterated LPPG is not available, making spectral assignment infeasible for apelin without isotopic enrichment.

Bottom Line: Type I beta-turns are initiated between R6 and L9, and a well-defined type IV beta-turn is initiated at S10.Furthermore, binding of apelin-17 to SDS micelles causes structuring of M15-F17, with no evidence for direct binding of this region to the micelles.These results are placed into the context of the membrane catalysis hypothesis for peptide-receptor binding, and a hypothetical mechanism of APJ binding and activation by apelin is advanced.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry & Molecular Biology, Dalhousie University, Halifax, Nova Scotia B3H 1X5, Canada.

ABSTRACT
Apelin is the peptidic ligand for the G-protein-coupled receptor APJ. The apelin-APJ system is important in cardiovascular regulation, fluid homeostasis, and angiogenesis, among other roles. In this study, we investigate interactions between apelin and membrane-mimetic micelles of the detergents sodium dodecyl sulfate (SDS), dodecylphosphocholine (DPC), and 1-palmitoyl-2-hydroxy-sn-glycero-3-[phospho-rac-(1-glycerol)] (LPPG). Far-ultraviolet circular dichroism spectropolarimetry and diffusion-ordered spectroscopy indicate that apelin peptides bind to micelles of the anionic detergents SDS and LPPG much more favorably than to zwitterionic DPC micelles. Nuclear magnetic resonance spectroscopy allowed full characterization of the interactions of apelin-17 with SDS micelles. Titration with paramagnetic agents and structural determination of apelin-17 in SDS indicate that R6-K12 is highly structured, with R6-L9 directly interacting with headgroups of the micelle. Type I beta-turns are initiated between R6 and L9, and a well-defined type IV beta-turn is initiated at S10. Furthermore, binding of apelin-17 to SDS micelles causes structuring of M15-F17, with no evidence for direct binding of this region to the micelles. These results are placed into the context of the membrane catalysis hypothesis for peptide-receptor binding, and a hypothetical mechanism of APJ binding and activation by apelin is advanced.

Show MeSH