Defining the Intramembrane Binding Mechanism of Sarcolipin to Calcium ATPase Using Solution NMR Spectroscopy

Jarrod J. Buffy; Bethany A. Buck-Koehntop; Fernando Porcelli; Nathaniel J. Traaseth; David D. Thomas; Gianluigi Veglia

doi:10.1016/j.jmb.2006.02.005

Defining the Intramembrane Binding Mechanism of Sarcolipin to Calcium ATPase Using Solution NMR Spectroscopy

Jarrod J. Buffy, Bethany A. Buck-Koehntop, Fernando Porcelli, Nathaniel J. Traaseth, David D. Thomas, Gianluigi Veglia

Chemical and Structural Biology (TMED)

Research output: Contribution to journal › Article › peer-review

45 Scopus citations

Abstract

Sarcolipin (SLN) is an integral membrane protein that is expressed in both skeletal and cardiac muscle, where it inhibits SERCA (calcium ATPase) by lowering its apparent Ca²⁺ affinity in a manner similar to that of its homologue phospholamban (PLN). We use solution NMR to map the structural changes occurring within SLN upon interaction with the regulatory target, SERCA, co-reconstituting the two proteins in dodecylphosphocholine (DPC) detergent micelles, a system that preserves the native structure of SLN and the activity of SERCA, with the goal of comparing these interactions with those of the previously studied PLN-SERCA complex. Our analysis of the structural dynamics of SLN in DPC micelles shows this polypeptide to be partitioned into four subdomains: a short unstructured N terminus (residues 1-6), a short dynamic helix (residues 7-14), a more rigid helix (residues 15-26), and an unstructured C terminus (residues 27-31). Upon addition of SERCA, the different domains behave according to their dynamics, molding onto the surface of the enzyme. Remarkably, each domain of SLN behaves in a manner similar to that of the corresponding domains in PLN, supporting the hypothesis that both SLN and PLN bind SERCA in the same groove and with similar mechanisms.

Original language	English (US)
Pages (from-to)	420-429
Number of pages	10
Journal	Journal of Molecular Biology
Volume	358
Issue number	2
DOIs	https://doi.org/10.1016/j.jmb.2006.02.005
State	Published - Apr 28 2006

Bibliographical note

Funding Information:
The authors graciously thank Professor Mac Lennan and Professor Toyoshima for the SERCA coordinates used for the model in Figure 6. This work was supported by National Institutes of Health grant (GM64742 and K02HL080081 to G.V. and GM27906 to D.D.T.) and American Heart Association grant 0160465Z. J.J.B. is supported by the Minnesota Craniofacial Research Training Program (MinnCResT), the NIH National Institute of Dental and Craniofacial Research (NIDCR 5T32-DE007288-10) and N.J.T. is supported by an American Heart Association Greater Midwest Affiliate Pre-Doctoral fellowship (0515491Z). NMR instrumentation at the University of Minnesota High Field NMR Center was funded by the National Science Foundation (BIR-961477) and the University of Minnesota Medical School.

Keywords

Ca-ATPase
SERCA
phospholamban
sarcolipin
solution NMR

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title = "Defining the Intramembrane Binding Mechanism of Sarcolipin to Calcium ATPase Using Solution NMR Spectroscopy",

abstract = "Sarcolipin (SLN) is an integral membrane protein that is expressed in both skeletal and cardiac muscle, where it inhibits SERCA (calcium ATPase) by lowering its apparent Ca2+ affinity in a manner similar to that of its homologue phospholamban (PLN). We use solution NMR to map the structural changes occurring within SLN upon interaction with the regulatory target, SERCA, co-reconstituting the two proteins in dodecylphosphocholine (DPC) detergent micelles, a system that preserves the native structure of SLN and the activity of SERCA, with the goal of comparing these interactions with those of the previously studied PLN-SERCA complex. Our analysis of the structural dynamics of SLN in DPC micelles shows this polypeptide to be partitioned into four subdomains: a short unstructured N terminus (residues 1-6), a short dynamic helix (residues 7-14), a more rigid helix (residues 15-26), and an unstructured C terminus (residues 27-31). Upon addition of SERCA, the different domains behave according to their dynamics, molding onto the surface of the enzyme. Remarkably, each domain of SLN behaves in a manner similar to that of the corresponding domains in PLN, supporting the hypothesis that both SLN and PLN bind SERCA in the same groove and with similar mechanisms.",

keywords = "Ca-ATPase, SERCA, phospholamban, sarcolipin, solution NMR",

author = "Buffy, {Jarrod J.} and Buck-Koehntop, {Bethany A.} and Fernando Porcelli and Traaseth, {Nathaniel J.} and Thomas, {David D.} and Gianluigi Veglia",

note = "Funding Information: The authors graciously thank Professor Mac Lennan and Professor Toyoshima for the SERCA coordinates used for the model in Figure 6. This work was supported by National Institutes of Health grant (GM64742 and K02HL080081 to G.V. and GM27906 to D.D.T.) and American Heart Association grant 0160465Z. J.J.B. is supported by the Minnesota Craniofacial Research Training Program (MinnCResT), the NIH National Institute of Dental and Craniofacial Research (NIDCR 5T32-DE007288-10) and N.J.T. is supported by an American Heart Association Greater Midwest Affiliate Pre-Doctoral fellowship (0515491Z). NMR instrumentation at the University of Minnesota High Field NMR Center was funded by the National Science Foundation (BIR-961477) and the University of Minnesota Medical School. ",

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doi = "10.1016/j.jmb.2006.02.005",

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T1 - Defining the Intramembrane Binding Mechanism of Sarcolipin to Calcium ATPase Using Solution NMR Spectroscopy

AU - Buffy, Jarrod J.

AU - Buck-Koehntop, Bethany A.

AU - Porcelli, Fernando

AU - Traaseth, Nathaniel J.

AU - Thomas, David D.

AU - Veglia, Gianluigi

N1 - Funding Information: The authors graciously thank Professor Mac Lennan and Professor Toyoshima for the SERCA coordinates used for the model in Figure 6. This work was supported by National Institutes of Health grant (GM64742 and K02HL080081 to G.V. and GM27906 to D.D.T.) and American Heart Association grant 0160465Z. J.J.B. is supported by the Minnesota Craniofacial Research Training Program (MinnCResT), the NIH National Institute of Dental and Craniofacial Research (NIDCR 5T32-DE007288-10) and N.J.T. is supported by an American Heart Association Greater Midwest Affiliate Pre-Doctoral fellowship (0515491Z). NMR instrumentation at the University of Minnesota High Field NMR Center was funded by the National Science Foundation (BIR-961477) and the University of Minnesota Medical School.

PY - 2006/4/28

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N2 - Sarcolipin (SLN) is an integral membrane protein that is expressed in both skeletal and cardiac muscle, where it inhibits SERCA (calcium ATPase) by lowering its apparent Ca2+ affinity in a manner similar to that of its homologue phospholamban (PLN). We use solution NMR to map the structural changes occurring within SLN upon interaction with the regulatory target, SERCA, co-reconstituting the two proteins in dodecylphosphocholine (DPC) detergent micelles, a system that preserves the native structure of SLN and the activity of SERCA, with the goal of comparing these interactions with those of the previously studied PLN-SERCA complex. Our analysis of the structural dynamics of SLN in DPC micelles shows this polypeptide to be partitioned into four subdomains: a short unstructured N terminus (residues 1-6), a short dynamic helix (residues 7-14), a more rigid helix (residues 15-26), and an unstructured C terminus (residues 27-31). Upon addition of SERCA, the different domains behave according to their dynamics, molding onto the surface of the enzyme. Remarkably, each domain of SLN behaves in a manner similar to that of the corresponding domains in PLN, supporting the hypothesis that both SLN and PLN bind SERCA in the same groove and with similar mechanisms.

AB - Sarcolipin (SLN) is an integral membrane protein that is expressed in both skeletal and cardiac muscle, where it inhibits SERCA (calcium ATPase) by lowering its apparent Ca2+ affinity in a manner similar to that of its homologue phospholamban (PLN). We use solution NMR to map the structural changes occurring within SLN upon interaction with the regulatory target, SERCA, co-reconstituting the two proteins in dodecylphosphocholine (DPC) detergent micelles, a system that preserves the native structure of SLN and the activity of SERCA, with the goal of comparing these interactions with those of the previously studied PLN-SERCA complex. Our analysis of the structural dynamics of SLN in DPC micelles shows this polypeptide to be partitioned into four subdomains: a short unstructured N terminus (residues 1-6), a short dynamic helix (residues 7-14), a more rigid helix (residues 15-26), and an unstructured C terminus (residues 27-31). Upon addition of SERCA, the different domains behave according to their dynamics, molding onto the surface of the enzyme. Remarkably, each domain of SLN behaves in a manner similar to that of the corresponding domains in PLN, supporting the hypothesis that both SLN and PLN bind SERCA in the same groove and with similar mechanisms.

KW - Ca-ATPase

KW - SERCA

KW - phospholamban

KW - sarcolipin

KW - solution NMR

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M3 - Article

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AN - SCOPUS:33646110094

SN - 0022-2836

VL - 358

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JF - Journal of Molecular Biology

IS - 2

ER -

Defining the Intramembrane Binding Mechanism of Sarcolipin to Calcium ATPase Using Solution NMR Spectroscopy

Abstract

Bibliographical note

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