title: BmSPN2, a serpin secreted by the filarial nematode Brugia malayi, does not inhibit human neutrophil proteinases but plays a noninhibitory role.
abstract: The filarial nematode, Brugia malayi, is a causative agent of lymphatic filariasis. Bm-spn-2, one of two serpin genes identified in B. malayi, is expressed only in humans where the encoded protein, BmSPN2, is secreted by blood-dwelling microfilariae. Previous work reported that BmSPN2 could inhibit the activities of elastase and cathepsin G from human neutrophils, despite an atypical amino acid sequence. This did not fit with accepted theories as to the sequence requirements of serpins for proteinase inhibition. We have cloned and expressed Bm-spn-2 in Escherichia coli and characterized the structural and functional properties of recombinant BmSPN2. Sequence alignment, circular dichroism spectroscopy, and susceptibility to cleavage by proteinases all suggest that BmSPN2 shares the tertiary structure typical of the serpin family including an accessible reactive center loop. However, we have found that BmSPN2 has no effect on the activity of neutrophil elastase or cathepsin G and does not form SDS-stable complexes with these proteinases. We provide evidence that BmSPN2 cannot undergo the characteristic stressed to relaxed transition required for proteinase inhibition by serpins. We conclude that BmSPN2 is not an atypical inhibitor but is a new noninhibitory serpin, in keeping with its sequence.
authors.affiliation: Division of Structural Medicine, Department of Haematology, Cambridge Institute for Medical Research, Wellcome Trust/MRC Building, Hills Road, United Kingdom.
authors.forename: Peter
authors.initials: P
authors.lastname: Stanley
authors.forename: Penelope E
authors.initials: PE
authors.lastname: Stein
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D017178:Brugia malayi / Q000235:genetics / Q000472:pathogenicity* / Q000502:physiology*; D056649:Cathepsin G; D002403:Cathepsins / Q000037:antagonists & inhibitors; D018509:DNA, Helminth / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics; D017238:Genes, Helminth; D015801:Helminth Proteins; D006801:Humans; D066298:In Vitro Techniques; D019272:Leukocyte Elastase / Q000037:antagonists & inhibitors; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009504:Neutrophils / Q000201:enzymology; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012697:Serine Endopeptidases; D015843:Serpins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism / Q000502:physiology*
chemical_list: C120921:Bm-SPN-2 protein, Brugia malayi; D018509:DNA, Helminth; D015801:Helminth Proteins; D011994:Recombinant Proteins; D015843:Serpins; D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; D019272:Leukocyte Elastase