The intramembrane active site of GlpG, an E. coli rhomboid protease, is accessible to water and hydrolyses an extramembrane peptide bond of substrates.
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Maegawa S, Koide K, Ito K, Akiyama Y
The intramembrane active site of GlpG, an E. coli rhomboid protease, is accessible to water and hydrolyses an extramembrane peptide bond of substrates.
Mol Microbiol. 2007 Apr;64(2):435-47.
- PubMed ID
- 17493126 [ View in PubMed]
- Abstract
Escherichia coli GlpG is an orthologue of the rhomboid proteases that catalyse intramembrane proteolysis of specific membrane proteins. We previously showed that it can cleave a type I model membrane protein, Bla-LY2-MBP, having the second transmembrane region of lactose permease (LY2) in vivo and in vitro at the predicted periplasm-membrane boundary region of LY2. Here we investigated the environment of the active site regions of GlpG in the membrane-integrated state by examining the modifiability of Cys residues introduced into the regions around the catalytic residues with membrane-permeable and -impermeable alkylating reagents. The results indicate that the enzyme active site is fully open to the external aqueous phase. GlpG also cleaved a similar fusion protein, Bla-GknTM-MBP, having the transmembrane region of Gurken (GknTM), a physiological substrate of Drosophila rhomboids. Engineered Cys residues in the cleavage site regions of the LY2 and GknTM sequences were efficiently modified with a membrane-impermeable alkylating reagent, showing that these regions are exposed to the periplasm. These results suggest that GlpG cleaves an extramembrane region of substrates, unlike the currently prevailing view that this class of membrane proteases acts against a membrane-embedded polypeptide segment after its lateral entrance into the enzyme's active site.