Human aspartic protease memapsin 2 cleaves the beta-secretase site of beta-amyloid precursor protein.
The cDNAs of two new human membrane-associated aspartic proteases, memapsin 1 and memapsin 2, have been cloned and sequenced. The deduced amino acid sequences show that each contains the typical pre, pro, and aspartic protease regions, but each also has a C-terminal extension of over 80 residues, which includes a single transmembrane domain and a C-terminal cytosolic domain. Memapsin 2 mRNA is abundant in human brain. The protease domain of memapsin 2 cDNA was expressed in Escherichia coli and was purified. Recombinant memapsin 2 specifically hydrolyzed peptides derived from the beta-secretase site of both the wild-type and Swedish mutant beta-amyloid precursor protein (APP) with over 60-fold increase of catalytic efficiency for the latter. Expression of APP and memapsin 2 in HeLa cells showed that memapsin 2 cleaved the beta-secretase site of APP intracellularly. These and other results suggest that memapsin 2 fits all of the criteria of beta-secretase, which catalyzes the rate-limiting step of the in vivo production of the beta-amyloid (Abeta) peptide leading to the progression of Alzheimer's disease. Recombinant memapsin 2 also cleaved a peptide derived from the processing site of presenilin 1, albeit with poor kinetic efficiency. Alignment of cleavage site sequences of peptides indicates that the specificity of memapsin 2 resides mainly at the S(1)' subsite, which prefers small side chains such as Ala, Ser, and Asp.
Lin
X
X
Protein Studies Program, Oklahoma Medical Research Foundation, and Department of Biochemistry, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA.
Koelsch
G
G
Wu
S
S
Downs
D
D
Dashti
A
A
Tang
J
J
D000544:Alzheimer Disease / Q000201:enzymology; D000595:Amino Acid Sequence; D053829:Amyloid Precursor Protein Secretases; D016564:Amyloid beta-Protein Precursor / Q000378:metabolism*; D016282:Aspartic Acid Endopeptidases / Q000737:chemistry* / Q000378:metabolism*; D001921:Brain / Q000201:enzymology; D003001:Cloning, Molecular; D010450:Endopeptidases; D004789:Enzyme Activation; D005455:Fluorescent Antibody Technique; D006367:HeLa Cells; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins; D016415:Sequence Alignment; D013379:Substrate Specificity
D016564:Amyloid beta-Protein Precursor; D012333:RNA, Messenger; D011994:Recombinant Proteins; D053829:Amyloid Precursor Protein Secretases; D010450:Endopeptidases; D016282:Aspartic Acid Endopeptidases; C509284:BACE2 protein, human; C509280:BACE1 protein, human