Post-translational secretion of fusion proteins in the halophilic archaea Haloferax volcanii.
Although protein secretion occurs post-translationally in bacteria and is mainly a cotranslational event in Eukarya, the relationship between the translation and translocation of secreted proteins in Archaea is not known. To address this question, the signal peptide-encoding region of the surface layer glycoprotein gene from the Haloarchaea Haloferax volcanii was fused either to the cellulose-binding domain of the Clostridium thermocellum cellulosome or to the cytoplasmic enzyme dihydrofolate reductase from H. volcanii. Signal peptide-cleaved mature versions of both the cellulose-binding domain and dihydrofolate reductase could be detected in the growth medium of transformed H. volcanii cells. Immunoblot analysis revealed, however, the presence of full-length signal peptide-bearing forms of both proteins inside the cytoplasm of the transformed cells. Proteinase accessibility assays confirmed that the presence of cell-associated signal peptide-bearing proteins was not due to medium contamination. Moreover, the pulse-radiolabeled signal peptide cellulose-binding domain chimera could be chased from the cytoplasm into the growth medium even following treatment with anisomycin, an antibiotic inhibitor of haloarchaeal protein translation. Thus, these results provide evidence that, in Archaea, at least some secreted proteins are first synthesized inside the cell and only then translocated across the plasma membrane into the medium.
Irihimovitch
Vered
V
Department of Life Sciences, Ben Gurion University of the Negev, Beersheva 84105, Israel.
Eichler
Jerry
J
D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry / Q000378:metabolism*; D001665:Binding Sites; D019622:Haloferax volcanii / Q000378:metabolism*; D008969:Molecular Sequence Data; D011498:Protein Precursors / Q000737:chemistry / Q000378:metabolism; D011499:Protein Processing, Post-Translational; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism*
D019843:Archaeal Proteins; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins