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Co‐reconstitution and Co‐crystallization of Phospholamban and Ca 2+ ‐ATPase a
Author(s) -
YOUNG HOWARD S.,
REDDY LAXMA G.,
JONES LARRY R.,
STOKES DAVID L.
Publication year - 1998
Publication title -
annals of the new york academy of sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.712
H-Index - 248
eISSN - 1749-6632
pISSN - 0077-8923
DOI - 10.1111/j.1749-6632.1998.tb08260.x
Subject(s) - phospholamban , transmembrane protein , atpase , chemistry , crystallography , transmembrane domain , biophysics , membrane , biochemistry , biology , phosphorylation , enzyme , receptor
Significant advances have recently been made in understanding the regulation of Ca 2+ ‐ATPase by phospholamban and in modeling their structures. However, these insights would be furthered by determining the 3‐D structure of both proteins within the membrane, thus revealing the structural basis for their interaction. To this end, we have developed methods for reconstituting purified Ca 2+ ‐ATPase with recombinant phospholamban. After reconstitution at high lipid‐to‐protein ratios, we have verified their functional association by measuring calcium transport and ATPase activity. Furthermore, we have grown co‐crystals after reconstitution at low lipid‐to‐protein ratios. The structure of Ca 2+ ‐ATPase has recently been solved by cryoelectron microscopy at 8‐Å resolution, thus revealing transmembrane α‐helices. Using a variety of constraints, we have associated these helices with the predicted transmembrane sequences to produce a detailed model for the packing of transmembrane helices. Structure determination of the co‐crystals is currently underway, which we hope will eventually reveal the interaction of phospholamban with Ca 2+ ‐ATPase at a similar level of detail.