Degradation of the amyloid beta-protein by the novel mitochondrial peptidasome, PreP.
Recently we have identified the novel mitochondrial peptidase responsible for degrading presequences and other short unstructured peptides in mitochondria, the presequence peptidase, which we named PreP peptidasome. In the present study we have identified and characterized the human PreP homologue, hPreP, in brain mitochondria, and we show its capacity to degrade the amyloid beta-protein (Abeta). PreP belongs to the pitrilysin oligopeptidase family M16C containing an inverted zinc-binding motif. We show that hPreP is localized to the mitochondrial matrix. In situ immuno-inactivation studies in human brain mitochondria using anti-hPreP antibodies showed complete inhibition of proteolytic activity against Abeta. We have cloned, overexpressed, and purified recombinant hPreP and its mutant with catalytic base Glu(78) in the inverted zinc-binding motif replaced by Gln. In vitro studies using recombinant hPreP and liquid chromatography nanospray tandem mass spectrometry revealed novel cleavage specificities against Abeta-(1-42), Abeta-(1-40), and Abeta Arctic, a protein that causes increased protofibril formation an early onset familial variant of Alzheimer disease. In contrast to insulin degrading enzyme, which is a functional analogue of hPreP, hPreP does not degrade insulin but does degrade insulin B-chain. Molecular modeling of hPreP based on the crystal structure at 2.1 A resolution of AtPreP allowed us to identify Cys(90) and Cys(527) that form disulfide bridges under oxidized conditions and might be involved in redox regulation of the enzyme. Degradation of the mitochondrial Abeta by hPreP may potentially be of importance in the pathology of Alzheimer disease.
Falkevall
Annelie
A
Department of Biochemistry and Biophysics, Stockholm University SE-106 91 Stockholm, Sweden.
Alikhani
Nyosha
N
Bhushan
Shashi
S
Pavlov
Pavel F
PF
Busch
Katrin
K
Johnson
Kenneth A
KA
Eneqvist
Therese
T
Tjernberg
Lars
L
Ankarcrona
Maria
M
Glaser
Elzbieta
E
D000368:Aged; D000369:Aged, 80 and over; D000595:Amino Acid Sequence; D016229:Amyloid beta-Peptides / Q000737:chemistry* / Q000378:metabolism; D000818:Animals; D002540:Cerebral Cortex / Q000473:pathology; D006801:Humans; D008297:Male; D008875:Middle Aged; D008928:Mitochondria / Q000378:metabolism*; D024101:Mitochondrial Proteins / Q000737:chemistry / Q000502:physiology*; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry*; D011485:Protein Binding; D051381:Rats; D017207:Rats, Sprague-Dawley; D017386:Sequence Homology, Amino Acid; D012697:Serine Endopeptidases / Q000737:chemistry / Q000502:physiology*; D013367:Submitochondrial Particles / Q000737:chemistry*
D016229:Amyloid beta-Peptides; D024101:Mitochondrial Proteins; D010455:Peptides; C514899:PREP protein, human; D012697:Serine Endopeptidases