The Pro Domain of β-Secretase Does Not Confer Strict Zymogen-like Properties but Does Assist Proper Folding of the Protease Domain
β-Secretase (BACE) is a membrane-bound aspartyl protease that cleaves the amyloid precursor protein to generate the N terminus of the amyloid β peptide. BACE is expressed as a precursor protein containing Pre, Pro, protease, transmembrane, and cytosolic domains. A soluble BACE derivative (PreProBA...
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Published in | The Journal of biological chemistry Vol. 276; no. 13; p. 10366 |
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Main Authors | , , , , , , , , , , , , , , |
Format | Journal Article |
Language | English |
Published |
American Society for Biochemistry and Molecular Biology
30.03.2001
|
Online Access | Get full text |
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Summary: | β-Secretase (BACE) is a membrane-bound aspartyl protease that cleaves the amyloid precursor protein to generate the N terminus
of the amyloid β peptide. BACE is expressed as a precursor protein containing Pre, Pro, protease, transmembrane, and cytosolic
domains. A soluble BACE derivative (PreProBACE460) that is truncated between the protease and transmembrane domains was produced
by baculovirus-mediated expression. ProBACE460 was purified from conditioned media of infected insect cells using immobilized
concanavalin A and immobilized BACE inhibitor, P10-P4â² Stat(Val). Furin cleaves ProBACE460 between the Pro and protease regions
to generate mature BACE460. The k
cat / K
m of ProBACE460 when assayed with a polypeptide substrate is only 2.3-fold less than that of BACE460. This finding and the
similar inhibitory potency of P10-P4â² Stat(Val) for ProBACE460 and BACE460 suggest that the Pro domain has little effect on
the BACE active site. Exposure of ProBACE460 to guanidine denaturation/renaturation results in a 7-fold higher recovery of
BACE activity than when BACE460 is similarly treated. The presence of free BACE Pro peptide during renaturation of BACE460
but not ProBACE460 increases recovery of activity. These findings show that the Pro domain in ProBACE460 does not suppress
activity as in a strict zymogen but does appear to facilitate proper folding of an active protease domain. |
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ISSN: | 0021-9258 1083-351X |
DOI: | 10.1074/jbc.M009200200 |