Degradation of tau protein by puromycin-sensitive aminopeptidase in vitro.
Tau, a microtubule associated protein, aggregates into intracellular paired helical filaments (PHFs) by an unknown mechanism in Alzheimer's disease (AD) and other tauopathies. A contributing factor may be a failure to metabolize free cytosolic tau within the neuron. The buildup of tau may then drive the aggregation process through mass action. Therefore, proteases that normally degrade tau are of great interest. A recent genetic screen identified puromycin-sensitive aminopeptidase (PSA) as a potent modifier of tau-induced pathology and suggested PSA as a possible tau-degrading enzyme. Here we have extended these observations using human recombinant PSA purified from Escherichia coli. The enzymatic activity and characteristics of the purified PSA were verified using chromogenic substrates, metal ions, and several specific and nonspecific protease inhibitors, including puromycin. PSA was shown to digest recombinant human full-length tau in vitro, and this activity was hindered by puromycin. The mechanism of amino terminal degradation of tau was confirmed using a novel N-terminal cleavage-specific tau antibody (Tau-C6g, specific for cleavage between residues 13-14) and a C-terminal cleavage-specific tau antibody (Tau-C3). Additionally, PSA was able to digest soluble tau purified from normal human brain to a greater extent than either soluble or PHF tau purified from AD brain, indicating that post-translational modifications and/or polymerization of tau may affect its digestion by PSA. These results are consistent with observations that PSA modulates tau levels in vivo and suggest that this enzyme may be involved in tau degradation in human brain.
Sengupta
Soma
S
Department of Cell and Molecular Biology, Feinberg School of Medicine at Northwestern University. s-sengupta@northwestern.edu
Horowitz
Peleg M
PM
Karsten
Stanislav L
SL
Jackson
George R
GR
Geschwind
Daniel H
DH
Fu
Yifan
Y
Berry
Robert W
RW
Binder
Lester I
LI
D000544:Alzheimer Disease / Q000201:enzymology; D000626:Aminopeptidases / Q000737:chemistry* / Q000235:genetics; D001921:Brain / Q000201:enzymology; D004789:Enzyme Activation; D006801:Humans; D011499:Protein Processing, Post-Translational; D011691:Puromycin / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D016875:tau Proteins / Q000737:chemistry* / Q000235:genetics
D011994:Recombinant Proteins; D016875:tau Proteins; D011691:Puromycin; D000626:Aminopeptidases; C025579:enkephalin degrading enzyme