Clustered hydrophobic amino acids in amphipathic helices mediate erlin1/2 complex assembly.
Erlin1 and erlin2 are highly homologous, ∼40kDa, endoplasmic reticulum membrane proteins that assemble into a ring-shaped complex with a mass of ∼2 MDa. How this complex is formed is not understood, but appears to involve multiple interactions, including a coiled-coil region that mediates lower-order erlin assembly, and a short hydrophobic region, termed the "assembly domain", that mediates higher-order assembly into ∼2 MDa complexes. Here we have used molecular modeling, mutagenesis and cross-linking to examine the role of the assembly domain in higher-order assembly. We find (i) that the assembly domains of erlin1 and erlin2 are amphipathic helices, (ii) that erlin1 alone and erlin2 alone can assemble into ∼2 MDa complexes, (iii) that higher-order assembly is strongly inhibited by point mutations to the assembly domain, (iv) that three interacting hydrophobic residues in the assembly domain and aromaticity are essential for higher-order assembly, and (iv) that while erlins1 and 2 are equally capable of forming lower-order homo- and hetero-oligomers, hetero-oligomers are the most prevalent form when erlin1 and erlin2 are co-expressed. Overall, we conclude that the ∼2 MDa erlin1/2 complex is composed of an assemblage of lower-order hetero-oligomers, probably heterotrimers, linked together by assembly domain hydrophobic residues.
Department of Pharmacology, SUNY Upstate Medical University, 750 E. Adams St., Syracuse, NY 13210, USA.
Deepa
D
Pednekar
Yuan
Y
Wang
Tatyana V
TV
Fedotova
Richard J H
RJ
Wojcikiewicz
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry* / Q000235:genetics; D000818:Animals; D006367:HeLa Cells; D006801:Humans; D057927:Hydrophobic and Hydrophilic Interactions; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics; D051379:Mice; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017433:Protein Structure, Secondary
D000596:Amino Acids; D008565:Membrane Proteins; C569220:erlin-1 protein, mouse; C569219:erlin-2 protein, mouse