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Mechanism of intramembrane proteolysis investigated with purified rhomboid proteases.
- Source :
-
The EMBO journal [EMBO J] 2005 Feb 09; Vol. 24 (3), pp. 464-72. Date of Electronic Publication: 2004 Dec 23. - Publication Year :
- 2005
-
Abstract
- Intramembrane proteases have the unusual property of cleaving peptide bonds within the lipid bilayer, an environment not obviously suited to a water-requiring hydrolysis reaction. These enzymes include site-2 protease, gamma-secretase/presenilin, signal peptide peptidase and the rhomboids, and they have a wide range of cellular functions. All have multiple transmembrane domains and, because of their high hydrophobicity, have been difficult to purify. We have now developed an in vitro assay to monitor rhomboid activity in the detergent solubilised state. This has allowed us to isolate for the first time a highly pure rhomboid with catalytic activity. Our results suggest that detergent-solubilised rhomboid activity mimics its activity in biological membranes in many aspects. Analysis of purified mutant proteins suggests that rhomboids use a serine protease catalytic dyad instead of the previously proposed triad. This analysis also suggests that other conserved residues participate in subsidiary functions like ligand binding and water supply. We identify a motif shared between rhomboids and the recently discovered derlins, which participate in translocation of misfolded membrane proteins.
- Subjects :
- Amino Acid Sequence
Animals
Bacillus subtilis enzymology
Bacillus subtilis genetics
Bacterial Proteins genetics
Bacterial Proteins isolation & purification
Bacterial Proteins metabolism
Detergents
Humans
In Vitro Techniques
Membranes enzymology
Models, Molecular
Molecular Sequence Data
Recombinant Proteins genetics
Recombinant Proteins isolation & purification
Recombinant Proteins metabolism
Sequence Homology, Amino Acid
Serine Endopeptidases genetics
Serine Endopeptidases isolation & purification
Solubility
Membrane Proteins metabolism
Serine Endopeptidases metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 0261-4189
- Volume :
- 24
- Issue :
- 3
- Database :
- MEDLINE
- Journal :
- The EMBO journal
- Publication Type :
- Academic Journal
- Accession number :
- 15616571
- Full Text :
- https://doi.org/10.1038/sj.emboj.7600537