Conformational properties of transmembrane polypeptide segments in the ER membrane

Abstract

We have developed a new experimental approach where the active site of oligosaccharyl transferase is used as a point of reference against which the position of a transmembrane segment in the membrane can be measured. This so-called glycosylation mapping technique allows any transmembrane segment to be positioned relative to a known reference transmembrane helix with known position in the membrane.

We have studied the position of transmembrane polypeptides in the ER membrane and determined the effects of single proline and charged residues on the position of a transmembrane helix in the endoplasmic reticulum membrane using the glycosylation mapping technique. We have found that proline residues can break a transmembrane helix when inserted near the end of the helix, but not when placed more centrally and only when the helix is sufficiently long. Compared to the helix-disrupting effects seen with proline residues, the charged amino acid residues cause less drastic changes. We suggest that charged residues do not break the helical conformation but just affect the position of the helix in the membrane.

The difference between the effects of positively and negatively charged residues can be explained by the so-called snorkel effect, i.e. that the very long side-chains of Arg and Lys can reach up along the transmembrane helix to allow the terminal charged moiety to reside in the lipid head group region. Our results from a turn propensity investigation suggest that in a very long transmembrane helix, twice as long as a normal helix, a single proline residue introduced near the center of the helix can induce the formation of two transmembrane segments separated by a tight turn.

The glycosylation mapping technique may be generally useful for determining the position of transmembrane helices in the membrane and the ends of different transmembrane segments.