Topological analysis of the lipoprotein organophosphate hydrolase from Sphingopyxis wildii reveals a periplasmic localisation
| Autor: | Hari Parapatla, Dayananda Siddavattam, Sunil Parthasarathy |
|---|---|
| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
Lipoproteins Topology Microbiology 03 medical and health sciences Bacterial Proteins Protein Domains Genetics Inner membrane Molecular Biology Diacylglycerol kinase biology Chemistry Periplasmic space Membrane transport biology.organism_classification Phosphoric Monoester Hydrolases Sphingomonadaceae Sphingopyxis Protein Transport Transmembrane domain 030104 developmental biology Biochemistry Membrane topology Periplasm Cysteine |
| Zdroj: | FEMS Microbiology Letters. 364 |
| ISSN: | 1574-6968 |
| Popis: | Organophosphate hydrolase (OPH) is a membrane-associated lipoprotein. It translocates across the inner membrane via the twin-arginine transport pathway and remains anchored to the periplasmic face of the inner membrane through a diacylglycerol moiety linked to the invariant cysteine residue found at the junction of a SpaseII cleavage site. Due to the existence of a transmembrane helix at the C-terminus of the mature OPH, an inner-membrane topology was predicted suggesting the C-terminus of OPH is cytoplasmic. The predicted topology was validated by generating OPH variants either fused in-frame with β-lactamase or with unique cysteine residues. Sphingopyxis wildii cells expressing OPH variants with Bla fused at the N-terminal, C-terminal or central regions all grew in the presence of ampicillin. Supporting the β-lactamase reporter assay, the OPH variants having unique cysteine residues at different strategic locations were accessible to the otherwise membrane-impermeant PEG-Mal (methoxypolyethylene glycol maleimide) revealing that, with the exception of the lipoprotein anchor, the entire OPH is in the periplasmic space. |
| Databáze: | OpenAIRE |
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