NDLI: Lipid bilayer stabilization of the Na,K-ATPase reconstituted in DPPC/DPPE liposomes

Content Provider	Springer Nature Link
Author	Rigos, Carolina Fortes Santos, Hérica de Lima Ward, Richard John Ciancaglini, Pietro
Copyright Year	2006
Abstract	Different subunit aggregates of the Na,K-ATPase may be formed depending on the method used to solubilize and purify the enzyme. We have studied the thermal unfolding of detergent-solubilized and dipalmitoylphosphatidylcholine/ dipalmitoylphosphatidylethanolamine liposome-reconstituted forms of the Na,K-ATPase by circular dichroism (CD) spectroscopy and p-nitrophenylphosphatase activity. The ellipticity at 222 nm of the solubilized and reconstituted forms showed a sigmoid decrease in the absolute value of the signal of 36 and 31% with T$_{50%}$ of 44 and 42°C, respectively. The catalytic activity was reduced in two steps with T$_{50%}$ of 32 and 52°C in the detergent-solubilized enzyme and T$_{50%}$ of 25 and 53°C in the reconstituted enzyme. The reduction in catalytic activity of the detergent-solubilized enzyme was bi-exponential with t $_{1/2}$ of 8.3 and 67.9 min, resulting in the total loss of activity after 120 min. However, under the same conditions, the ATPase activity of the reconstituted enzyme was reduced by approx 35% with a t $_{1/2}$ of 145 min. The results suggest that the α-and β-subunits present different thermal stability that may be modulated by the nature of the co-solvent (detergent or lipid) used in the preparations of the Na,K-ATPase. In addition, distinct processes of β-subunit displacement and α-α-subunit aggregate formation may also contribute to the changes in both the CD spectra and the enzyme activity. Furthermore, we have demonstrated the protective role of the phospholipid bilayer in the reconstituted enzyme compared with the detergent-solubilized enzyme.
Starting Page	438
Ending Page	445
Page Count	8
File Format	PDF
ISSN	10859195
Journal	Cell Biochemistry and Biophysics
Volume Number	44
Issue Number	3
e-ISSN	15590283
Language	English
Publisher	Humana Press
Publisher Date	2006-01-01
Publisher Place	Totowa
Access Restriction	One Nation One Subscription (ONOS)
Subject Keyword	Biochemistry Pharmacology/Toxicology Biotechnology Cell Biology
Content Type	Text
Resource Type	Article
Subject	Cell Biology Medicine Biochemistry Biophysics

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