Title

The Inhibitory Action Of Phospholamban Involves Stabilization Of Α-Helices Within The Ca-Atpase

Abstract

We have used attenuated total reflection Fourier transform infrared (ATR-FTIR) and circular dichroism (CD) spectroscopies to identify secondary and dynamic structural changes within the Ca-ATPase that result from the functional inhibition of transport activity by phospholamban (PLB). Isotopically labeled [13C]PLB was expressed and purified from Escherichia coli and was functionally reconstituted with unlabeled Ca-ATPase, permitting the resolution of the amide I and II absorbance bands of the Ca-ATPase from those of [13C]PLB. Upon co-reconstitution of the Ca-ATPase with PLB, spectral shifts are observed in both the CD spectra and the amide I and II bands associated with the Ca-ATPase, which are indicative of increased α-helical stability. Corresponding changes in the kinetics of H/D exchange occur upon association with PLB, indicating that 100 ± 20 residues in the Ca-ATPase that normally undergo rapid amide H/D exchange become exchange resistant. There are no corresponding large changes in the secondary structure of PLB. The affinity of the structural interaction between PLB and the Ca-ATPase is virtually identical to that associated with functional inhibition (Kd = 140 ± 30, μM), confirming that the inhibitory regulation of the Ca-ATPase by PLB involves the stabilization of α-helices within the Ca-ATPase.

Publication Date

1-22-2002

Publication Title

Biochemistry

Volume

41

Issue

3

Number of Pages

741-751

Document Type

Article

Personal Identifier

scopus

DOI Link

https://doi.org/10.1021/bi011148d

Socpus ID

0037154095 (Scopus)

Source API URL

https://api.elsevier.com/content/abstract/scopus_id/0037154095

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