Predicting the orientation of eukaryotic membrane-spanning proteins

We have developed a rule to predict the orientation of the first internal signal-anchor sequence in eukaryotic transmembrane proteins synthesized on the rough endoplasmic reticulum. The difference in the charges of the 15 residues flanking the first internal signal-anchor determine its orientation, with the more positive portion facing the cytosol. In proteins that span the membrane more than once, the orientation of all subsequent transmembrane segments would be determined by that of the most N-terminal one.