Stability and Global Fold of the Mouse Prohormone Convertase 1 Pro-Domain

Publisher Website
Google Scholar
Abstract
We have purified the mouse prohormone convertase 1 (PC1) pro-domain expressed in Escherichia coli cells and demonstrated, using a number of biophysical methods, that this domain is an independent folding unit with a Tm of 39 °C at a protein concentration of 20 μM and pH 7.0. This differs significantly from similar pro-domains in bacteria and human furin, which are unfolded at 25 °C and require the catalytic domain in order to be structured [Bryan et al. (1995) Biochemistry 34, 10310−10318; Bhattacharjya et al. (2000) J. Biomol. NMR 16, 275−276]. Using heteronuclear NMR spectroscopy, we have determined the backbone 1H, 13C, and 15N assignments for the pro-domain of PC1. On the basis of 1H/13C chemical shift indices, NOE analysis, and hydrogen exchange measurements, the pro-domain is shown to consist of a four-stranded β-sheet and two α-helices. The results presented here show that both the bacterial pro-domain in complex with subtilisin and the uncomplexed mouse PC1 pro-domain have very similar overall folds despite a lack of sequence homology. The structural data help to explain the location of the secondary processing sites in the pro-domains of the PC family, and a consensus sequence for binding to the catalytic domain is proposed.