Use of a new label, ¹³C = ¹⁸O, in the determination of a structural model of phospholamban in a lipid bilayer. Spatial restraints resolve the ambiguity arising from interpretations of mutagenesis data

A structural model of pentameric phospholamban (Plb) in a lipid bilayer has been derived using a combination of experimental data, obtained from ATR-FTIR site-directed dichroism, and the implementation of the resulting restraints during a molecular dynamics simulation. Plb (residues 24-52) has been synthesised incorporating a new label, 1-¹³C = ¹⁸O, at residues 42 and 43. We have not only determined the tilt of the helices, 10(±6)°, but also the relative orientation of the transmembrane segments, with an ω angle of -32(±10)°for L42. This angle is taken as zero in the direction of the helix tilt. Plb is a simple test case where site-directed dichroism has been applied to resolve the indeterminacy arising from the mutagenesis data available. The results presented point specifically to a single structural model for Plb. (C) 2000 Academic Press.