Lecture 23 Apr 2007

General Principles of Membrane Protein Folding and Stability from the lab of Stephen White, UC Irvine

Membrane proteins of known structure

Membrane proteins

It is difficult to crystallize membrane proteins, because they require detergents for solubilization. To crystallize, the detergent must be reduced without leading to non-specific aggregation. There are only 124 unique proteins in the PDB currently. The pace of new structures is exponential, but greatly lags the expansion of the overall PDB.

White SH
The progress of membrane protein structure determination.
Protein Sci. 2004 Jul;13(7):1948-9.

Figure

The typical membrane is 40-60Å across. It includes a central, nonpolar region of 30Å filled by fatty acyl chains of the phospholipids, and two more polar regions at each surface. (Fig. 1-29)

Most membrane proteins have transmembrane domains of bundles of alpha-helices (Fig. 1-30) or antiparallel beta-barrels (Fig. 1-32). (Jmol-8 strands) (Jmol-12 strands)

There are also membrane proteins, called monotopic, which are held to one surface of a membrane by helices that enter only one leaflet of the bilayer. (Jmol)

By sequence analysis it is fairly easy to detect the regions of 20 consecutive amino acids of nonpolar character , which are necessary, in general, to span a lipid bilayer (Fig. 1-31). PredictProtein (use HTM) and other servers can also be used to do this. It is more difficult to predict the transmembrane regions of β-sheet membrane proteins.

This protein has 120 amino acids (the image was accidently clipped). Three transmembrane helices are predicted; about 10-25, 30-45 and 80-100. The orientation is also predicted: the C terminus is on the inside (cytoplasm of the bacterium) and the N-terminus is outside (periplasm). This follows the positive-inside rule, in which the net charge of inside loops tend to be positive. Notice the two Arg around 25 and the 3 around 105.

An interesting feature of this protein is that two of the transmembrane regions are predicted to contain acidic groups: Glu 36 and Glu 92. These are likely to be important for structure or function.

In addition to the nonpolar amino acids that populate the transmembrane spans: ALIVMF, other residues can be found also:

NSTC are fairly common, as they can make H-bonds to the backbones of other helices.
P is also fairly common in membrane helices, for reasons that are not yet clear.
G is found at close contact sites between helices.
WY are often found at the lipid-aqueous interface, due to the amphipathic nature of their side chains.

This protein is found in the "purple membranes" of halophilic bacteria. It has 248 amino acids and it was known from early EM work by Richard Henderson at Cambridge UK that it has seven α-helical transmembrane spans. It functions as a light-driven proton pump. It pumps protons out, so that it builds a conventional proton gradient that can be used for ATP synthesis, or coupled ion transport. It absorbs light through a retinal group that is covalently attached to Lys 216. (Jmol)

Water, glycerol and ammonia all can be transported across biological membranes by members of a family of transport proteins. These transporters appear in all branches of life. Humans have at least 10 such genes. They can be recognized by a signature sequence, and an overall duplication. A curious feature is that the two, related halves of the protein have opposite orientation in the membrane. A second unusual feature in the case of aquaporins is that a conserved NPA sequence is part of a half-spanning segment. A loop extends halfway through the bilayer and then returns to the original surface as an α-helix. The 2 asparagines interact at the middle of the membrane. This helps to form the very tight pathway for the particular molecule that is transported. Furthermore, these transporters form tetramers of C4 symmetry, and the central channel is an ion channel, in some cases. (Jmol of a glycerolporin)