A proprotein convertase-inhibiting serpin with an endoplasmic reticulum targeting signal from Branchiostoma lanceolatum, a close relative of vertebrates.
Lancelets are considered to take a key position in the evolution of lineages leading to vertebrates. Herein, a serpin from the lancelet Branchiostoma lanceolatum, Bl-Spn1, was identified that inhibits the PCs (proprotein convertases) PC1/3 and furin. The inhibitor forms SDS-stable complexes with either of its targets. Analysis of the inhibitor/furin reaction products by mass spectroscopy assigns the enzyme's cleavage position C-terminally to Met-Met-Lys-Arg downward arrow in the reactive site loop of Spn1, in concordance with the classical recognition/cleavage site of the principal vertebrate PCs. The inhibitor is equipped with a canonical ER (endoplasmic reticulum) retrieval signal, Lys-Asp-Glu-Leu (KDEL), marking the inhibitor as a guardian of the cellular secretory routes. Deletion of the ER retrieval signal results in the export of the inhibitor into the medium of transfected COS-7 cells, consistent with the assigned intracellular location. These results identify Bl-Spn1 as the first serpin that may inhibit PC1/3-like subtilases at their natural sites of action. Phylogenetic comparisons support a concept implying a general role for ER-residing serpins in the surveillance of subtilase-like enzymes along the constitutive and regulated secretory pathways of metazoans including a role in the defence of intruders that turn PCs to their propagation.
Bentele
Caterina
C
Department of Biotechnology, Faculty of Technology, University of Bielefeld, Bielefeld D-33501, Germany.
Krüger
Olaf
O
Tödtmann
Ulf
U
Oley
Mareke
M
Ragg
Hermann
H
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002816:Chordata, Nonvertebrate / Q000235:genetics / Q000378:metabolism*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D005091:Exons / Q000235:genetics; D045683:Furin / Q000737:chemistry / Q000378:metabolism; D007438:Introns / Q000235:genetics; D013058:Mass Spectrometry; D008969:Molecular Sequence Data; D043484:Proprotein Convertases / Q000037:antagonists & inhibitors* / Q000378:metabolism*; D011485:Protein Binding; D046188:Protein Modification, Translational; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012697:Serine Endopeptidases / Q000378:metabolism; D015842:Serine Proteinase Inhibitors / Q000378:metabolism; D015843:Serpins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D012967:Sodium Dodecyl Sulfate / Q000494:pharmacology
D018076:DNA, Complementary; D011994:Recombinant Proteins; D015842:Serine Proteinase Inhibitors; D015843:Serpins; D012967:Sodium Dodecyl Sulfate; D043484:Proprotein Convertases; D012697:Serine Endopeptidases; D045683:Furin