The C-terminus of prenylin is important in forming a dimer conformation necessary for endoplasmic-reticulum-to-Golgi transport.
Prenylin [or prenylated Rab acceptor 1 (PRA1)] is a multi-pass transmembrane protein that initially inserts into the ER (endoplasmic reticulum) membrane, followed by vesicular transport along the exocytic pathway to the Golgi complex where it may regulate the functions of prenylated proteins. Deletion of the C-terminal 10 amino acid residues of prenylin blocks its export from the ER. We have employed site-directed mutagenesis to investigate the role of each of the C-terminal 10 residues in the ER export of prenylin. This region contains a di-acidic motif (Asp176-Xaa-Glu), but changing either acidic residue to alanine has no effect on the ER export of prenylin. Alanine-scanning mutagenesis of the entire C-terminal region reveals that only the very C-terminal Val185 residue is crucial for the ER export of prenylin. Changing the C-terminal Val185 to most other amino acids effectively prevents prenylin from exiting the ER. However, deletion of Val185 has only moderate effect on the ER export of prenylin, suggesting that this valine residue is not part of an export signal itself; instead, it may affect the folding and conformation of prenylin. We show that the wild-type prenylin can efficiently form a homodimer in the cell by using a cell-permeant cross-linker, whereas the large C-terminal truncation and Val185 mutants are defective in forming such a dimer. Thus we have identified a single C-terminal valine residue that is essential for the proper dimerization and ER export of prenylin.
Liang
Zhimin
Z
Department of Biochemistry and Molecular Biology, University of Oklahoma Health Sciences Center, 940 S.L. Young Blvd., BMSB 853, Oklahoma City, OK 73104, USA.
Veeraprame
Helga
H
Bayan
Nami
N
Li
Guangpu
G
D020816:Amino Acid Motifs; D000818:Animals; D002460:Cell Line / Q000378:metabolism; D006224:Cricetinae; D003432:Cross-Linking Reagents / Q000494:pharmacology; D019281:Dimerization; D018871:Endoplasmic Reticulum, Smooth / Q000378:metabolism*; D006056:Golgi Apparatus / Q000378:metabolism*; D007668:Kidney; D008565:Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D008647:Mesocricetus; D051379:Mice; D008954:Models, Biological; D016297:Mutagenesis, Site-Directed; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D017384:Sequence Deletion; D013329:Structure-Activity Relationship; D014162:Transfection; D014633:Valine / Q000737:chemistry
D003432:Cross-Linking Reagents; D008565:Membrane Proteins; D011993:Recombinant Fusion Proteins; C492518:prenylin protein, mouse; D014633:Valine