(6) However, such chaperones remain unidentified in prokaryotes. In eukaryotes, ribosome-associating proteins serve as chaperones for tail-anchored membrane proteins. (5) Since their tails are too short, they are released from the ribosomes after synthesis and cannot use the SRP/SR and Sec translocons. These proteins possess one or two transmembrane regions connected with a short C-terminal tail. Some small membrane proteins called Sec-independent proteins do not require SRP and Sec translocons for integration ( Figure 1c). The nascent protein requires MPIase to integrate into the membrane, and YidC accelerates the integration in concert with MPIase. (c) Some small membrane proteins are integrated into the membrane without using a Sec translocon (translocon-independent). The signal sequence of the preprotein is cleaved using signal peptidase at the periplasm, and the secretory protein is released. The motor protein SecA hydrolyzes ATP as it catalyzes the protein insertion. The preprotein with a signal sequence is targeted to a protein complex consisting of SecYEG, SecDF(yajC), and YidC. (b) After the synthesis of the secretory protein (preprotein) in the ribosome, the molecular chaperone SecB binds the preprotein. coli, SRP comprises Ffh and 4.5S RNA, and SR corresponds to FtsY. YidC can function as a catalyst for membrane integration and facilitate the lateral movement of transmembrane segments into the bilayers. The growing peptide chain threading from the ribosome into the translocon is extruded into the lipid bilayer according to the conformational change of the translocon. The ribosome–nascent chain is further transferred to SecYEG. (a) In the translocon-dependent membrane protein integration pathway, the signal recognition particle (SRP) recognizes the signal peptide generated from a ribosome and leads it to the SRP receptor (SR). MPIase enhances the function of SecYEG and YidC however, colocalization of MPIase with these factors is speculative at this stage. Schematic model for membrane protein integration and translocation in E. (3,4) Also, SecYEG is used to translocate a protein across the membrane ( Figure 1b). Then, the ribosome–nascent protein complex is delivered to a channel complex called the SecYEG translocon and a membrane protein insertase termed YidC. (2) The signal recognition particle (SRP) binds the hydrophobic sequence from a ribosome to an SRP receptor (SR) on the membrane. coli), protein integration into the membrane is triggered by recognizing the transmembrane region of a nascent protein ( Figure 1a). (1) In the inner membrane of Escherichia coli ( E. These processes require the assembly of proteins, called translocons, that are widely conserved from prokaryotes to higher eukaryotes.

Therefore, integrating membrane proteins into the membrane and translocating secretory proteins across the membrane in conjunction with protein synthesis are extremely essential. After their synthesis according to genetic information, they are transported to the proper place where they work. Proteins are indispensable constituents for the survival of all living organisms. These results demonstrated the direct interactions of MPIase with a substrate protein and revealed detailed mechanisms of membrane protein integration. The high affinity of MPIase to the hydrophobic region and the basic amino acid residues of the protein was suggested by docking simulations and proven experimentally by SPR using protein mutants devoid of target regions. These experiments yielded that the 6- O-acetyl group on glucosamine and the phosphate of MPIase play important roles leading to interactions with the Pf3 coat protein. To focus on more detailed MPIase substructures, we performed docking simulations and saturation transfer difference-nuclear magnetic resonance. Compared with mini-MPIase-3, natural MPIase showed faster association and dissociation due to its long sugar chain despite the slight difference in affinity.

Surface plasmon resonance (SPR) analyses revealed the importance of a pyrophosphate for affinity to the Pf3 coat protein. Here, to elucidate integration mechanisms using MPIase, we analyzed intermolecular interactions of MPIase or its synthetic analogs with a model substrate, the Pf3 coat protein, using physicochemical methods.

We reported that a synthesized minimal unit of MPIase possessing only one trisaccharide, mini-MPIase-3, involves an essential structure for the integration activity. Additionally, a glycolipid named MPIase (membrane protein integrase), composed of a long sugar chain and pyrophospholipid, was proven essential for membrane protein integration. Integration of proteins into biomembranes in Escherichia coli is mediated by proteinaceous factors, such as Sec translocons and an insertase YidC. Inducing newly synthesized proteins to appropriate locations is an indispensable biological function in every organism.

Membrane pro llc