[ccp4bb] off topic: GPCR membane insertion/orientation

[Justin Hall]

Dear Community,

In trying to trouble shoot an experiment I have become interested in  
the cellular process that regulates the insertion and proper  
orientation of membrane proteins. I am looking for references for how  
a GPCR is correctly oriented during expression (i.e. the extra  
cellular domain ends up extra cellularly oriented instead of a 50/50  
mix in and out), my intuition is that there must be an N-terminal  
sequence that directs this process, but I am having no luck finding  
information on what this sequence is for GPCRs, what players are  
involved or how orientation is thought to be controlled. Any  
suggestions?

This is all spurred by my wanting to use phage display with a protein  
that binds to the intracellular side of a GPCR, but of course that is  
the hard side to present to the outside of a cell so I need to figure  
out how to flip these guys around. I have thought about adding a new  
TM helix before TM1 (or removing TM1) to flip these guys, but was  
hoping there might be another way around that doesn't involve such  
massive architectural rearrangement such as simply clipping the  
N-terminal sequence responsible for proper orientation (if such a  
thing exists). Cheers~

~Justin