Molecular cloning of endopin 1, a novel serpin localized to neurosecretoryvesicles of chromaffin cells - Inhibition of basic residue-cleaving proteases by endopin 1
Sr. Hwang et al., Molecular cloning of endopin 1, a novel serpin localized to neurosecretoryvesicles of chromaffin cells - Inhibition of basic residue-cleaving proteases by endopin 1, J BIOL CHEM, 274(48), 1999, pp. 34164-34173
Serpins represent a diverse class of endogenous protease inhibitors that re
gulate important biological functions. In consideration of the importance o
f regulated proteolysis within secretory vesicles for the production of pep
tide hormones and neurotransmitters, this study revealed the molecular iden
tity of a novel serpin, endopin 1, that is localized to neurosecretory vesi
cles of neuropeptide-containing chromaffin cells (chromaffin granules). End
opin 1 of 68-70 kDa was present within isolated chromaffin granules. Stimul
ated cosecretion of endopin 1 with chromaffin granule components, [Met]enke
phalin and a cysteine protease known as "pro-hormone thiol protease," demon
strated localization of endopin 1 to functional secretory vesicles. Punctat
e, discrete immunofluorescence cellular localization of endopin 1 in chroma
ffin cells was consistent with its secretory vesicle localization. Endopin
1 contains a unique reactive site loop with Arg as the predicted P1 residue
, suggesting inhibition of basic residue-cleaving proteases; indeed, trypsi
n was potently inhibited (K7(i(app)) of 5 nM), and plasmin was moderately i
nhibited. Although endopin 1 possesses homology with alpha(1)-antichymotryp
sin, chymotrypsin was not inhibited. Moreover, endopin 1 inhibited the chro
maffin granule prohormone thiol protease (involved in proenkephalin process
ing), These results suggest a role for the novel serpin, endopin 1, in regu
lating basic residue-cleaving proteases within neurosecretory vesicles of c
hromaffin cells.