Bio- chemical analyses demonstrate PAT complex engages nascent TMDs that contain unshielded polar amino acids but disengages upon substrate folding. A diverse set of multipass membrane proteins show impaired biogenesis upon PAT complex depletion, despite correct topological insertion. The PAT complex is an obligate heterodimer consisting of the highly conserved proteins CCDC47 and Asterix. Following accurate insertion of TMD1, we define the engagement of a newly identified intramembrane chaperone protein complex that we term the PAT complex. Finally, EMC necessity could be by- passed by enforcement of TMD1 topology via an N-terminal signal peptide. EMC and SRP receptor were sufficient for the correct insertion of TMD1, while insertion of the next TMD required Sec61. In the absence of EMC, TMD1 was topologically inverted or failed to insert altogether. This is supported by biochemical reconstitution of β1-adrenergic receptor (β1AR) insertion in-vitro, which placed EMC at an early step during co-translational insertion of the first TMD (TMD1). Neither the mechanisms that facilitate accurate topological insertion or the subsequent stabilization of polar TMDs during IMP biogenesis are completely understood.įirst, we demonstrate the efficient topogenesis of many GPCRs requires the conserved ER membrane protein complex (EMC). Productive bio- genesis of most “multipass” membrane proteins requires insertion in a defined topology as well as packing of their TMDs into a helical bundle, often through inter- actions between polar residues unstably located in the hydrophobic lipid bilayer. While some contain a single TMD, most feature many biophysically unique TMDs. This thesis aims to describe our recent contributions to the understanding of how membrane proteins are made at the endoplasmic reticulum (ER), the primary site of IMP biogenesis in the cell.Īlthough IMPs are defined by a single feature, a transmembrane domain (TMD), the ~5000 encoded in the mammalian genome are diverse. Highlighting their importance is a myriad of human diseases observed upon disruption of their biogenesis. All these processes require integral membrane proteins (IMPs), which are embedded within all cellular membranes. However, cells do not live in isolation and must communicate with neighboring cells, uptake and traffic nutrients, and react to a dynamic extracellular environment. Semi-permeable membranes facilitate autonomy via an outer barrier (plasma membrane) and enclose functional hubs (organelles) to efficiently carry out biological processes. A crucial aspect of cellular physiology is the ability of a single cell to remain autonomous and concentrate reagents to improve efficiency.
0 kommentar(er)
