|Cytoplasmic membrane proteins were identified by using the maxH algorithm.|
Ref.: Boyd, Schierle and Beckwith (1998) How Many Membrane Proteins Are There? Protein Science 7(1):201-205.
Subsequently, the topology of cytoplasmic membrane proteins was predicted using the TMHMM server version 2.0. These predictions were then compared with the topological model of the relevant SwissProt entries. One model was chosen based on plausibility considerations. These models often include changes in the start and end points of transmembrane segments (TM). For example, it makes little sense to start or end a TM with a charged residue, which is often ignored by computational predictions.
Protein sequences are presented in lower case, the TMs in upper case.
Some rules of thumb for topology predictions
The positive inside rule postulates that cytoplasmic segments are containing significantly more positive charges than extracytoplasmic (periplasmic) segments.
This rule helps to
1. predict the cellular localisation of N- and C-termini of the membrane protein
2. predict if there is an even or odd number of TMs
3. to decide if an uncertain TM is real (by checking if the charge distribution in loops follows the positive inside rule).
TMs are often flanked by aromatic residues that help the membrane protein to be more stably embedded within the lipid bilayer.
If in doubt or to improve the prediction, an alignment of family members will often be able to indicate if a TM is real or not. In these alignments, do not allow gaps to occur within TMs. Because TMs are helices, each addition or deletion of a single residue will cause rotation of the helix faces. This may not be critical for proteins containing single TMs. However, polytopic membrane proteins do usually not tolerate such manipulations because interaction with other TMs can no longer occur productively if helix faces are altered.
Topology prediction online
TMHMM server version 2.0
ISREC mss pred Server
CBS prediction servers
Positive inside rule
Determinants of membrane protein topology Boyd D, Manoil C, Beckwith J 1987
Control of topology and mode of assembly of a polytopic membrane protein by positively charged residues. von Heijne G. 1989
Translocation of N-terminal tails across the plasma membrane. Cao G, Dalbey RE. 1994
Sec-independent translocation of a 100-residue periplasmic N-terminal tail in the E. coli inner membrane protein proW. Whitley P, Zander T, Ehrmann M, Haardt M, Bremer E, von Heijne G. 1994
phoA fusions, a genetic method to test topological models
A genetic approach to analyzing membrane protein topology. Manoil C, Beckwith J. 1986
Genetic analysis of membrane protein topology by a sandwich gene fusion
approach. Ehrmann M, Boyd D, Beckwith J. 1990