Effects of Spin-Labels on Membrane Burial Depth of MARCKS-ED Residues

Site-directed spin-labeling electron paramagnetic resonance spectroscopy is a useful tool to obtain information about the environment of specific residues. One of its applications is to investigate membrane protein topology based on the accessibility of the spin label, with the assumption that the position of the spin label in the membrane is close to that of the native residue. This assumption is valid in proteins with well-ordered structures, but could be problematic in small peptides because the labeling may cause a perturbation that is large enough to change local interactions between the peptide and the membrane. To quantitatively characterize such effects, we have simulated the association of a 25-amino-acid peptide, MARCKS-ED, to membranes with and without spin labels. Our simulations show that the depths of spin labels are ∼6-17 Å deeper than the unlabeled charged and polar residues in the wild-type. When the hydrophobic residue Phe is labeled, however, the spin-label depth is close to that of the native residue as well as the experimental value. Our study suggests that one should be cautious in interpretation of spin label data when charged and polar residues in small peptides are labeled.