Detection of a soluble form of BACE-1 in human cerebrospinal fluid by a sensitive activity assay.
BACKGROUND
Formation of deposits of the insoluble amyloid beta-peptide is believed to be causally related with neurodegeneration in Alzheimer disease (AD). The beta-peptide originates from a larger amyloid precursor protein (APP) by the action of proteolytic enzymes. The first proteolytic event leading to amyloid formation is the cleavage of APP by the membrane-bound aspartyl protease BACE-1, also known as memapsin-2. Inhibition of BACE-1 is thought to be a therapeutic approach to AD. Measuring BACE-1 activity in biological samples would be useful to elucidate the mechanism of AD and for development of AD drugs.


METHODS
We developed a sensitive and specific activity assay for BACE-1. The assay is based on a genetically engineered proenzyme that is specifically activated by BACE-1. The resulting active enzyme is measured with a chromogenic substrate. The use of 2 coupled reactions produces a detection limit as low as 0.4 pmol/L.


RESULTS
The assay detected BACE-1 activity in extracts of human brain tissue as well as, unexpectedly, in human cerebrospinal fluid (CSF). Gel electrophoresis and Western blotting identified the BACE-1 present in CSF as a truncated soluble form of the originally membrane-bound BACE-1.


CONCLUSIONS
Detection of the soluble form of BACE-1 in CSF, a relatively easily accessible biological fluid, may be useful for monitoring the effects of drug candidates in vivo and may have diagnostic or prognostic applications.
Verheijen
Jan H
JH
TNO Quality of Life, Biomedical Research, Leiden, The Netherlands. JH.Verheijen@pg.tno.nl
Huisman
Linda G M
LG
van Lent
Natascha
N
Neumann
Ulf
U
Paganetti
Paolo
P
Hack
C Erik
CE
Bouwman
Femke
F
Lindeman
Jan
J
Bollen
Edward L E M
EL
Hanemaaijer
Roeland
R
D053829:Amyloid Precursor Protein Secretases; D016282:Aspartic Acid Endopeptidases; D001921:Brain / Q000201:enzymology; D053148:Caspase 3; D020169:Caspases / Q000737:chemistry; D003429:Cross Reactions; D010450:Endopeptidases / Q000134:cerebrospinal fluid* / Q000737:chemistry; D006801:Humans; D007118:Immunoassay; D011498:Protein Precursors / Q000737:chemistry; D012680:Sensitivity and Specificity; D012995:Solubility; D014020:Tissue Extracts
D011498:Protein Precursors; D014020:Tissue Extracts; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases; D016282:Aspartic Acid Endopeptidases; C509280:BACE1 protein, human