The increased accumulation of A4 (or beta)_ amyloid in senile plaques and around blood vessels appears to be an early and distinctive feature of Alzheimer's disease. A4 amyloid is peptide of at most 43 amino which is produced by proteolytic processing of an amyloid precursor protein, a sulfated proteoglycan abundantly synthesized in normal brain, and in other tissues. In Alzheimer's disease proteolytic processing of this amyloid precursor protein appears disordered to produce increased amounts of the A4 amyloid peptide. The gene for amyloid precursor protein has been cloned and the amino sequence deduced. Therefore, the peptide bands hydrolyzed by endoproteases to produce the amino and carboxy termini of A4 amyloid can be surmised. Based on the known properties of endoproteases and our preliminary data presented here, we believe that metalloendoprotease in brain can cleave the amyloid precursor protein to produce the carboxy terminus of A4 amyloid. Based on our preliminary data, presented the custom synthesized fluorogenic endoprotease substrate Methoxy-Succinyl-Thr-Val-Ile-4 amino methyl coumarin which was synthesized with the peptide sequence in the amyloid precursor protein hydrolyzed to produce the carboxy terminus of A4 amyloid. We have already shown that this brain endoprotease requires zinc for catalytic activity and is inhibited by three specific active site reversible metalloendoprotease inhibitors.
The specific aims of this pilot project are to: 1. Further characterize the biochemistry of the brain metalloendoprotease which hydrolyzes the synthetic amyloid substrate Me Succ-Thr-Val-lie-AMC. 2. Determine whether this metalloendoprotease is different in its biochemical properties, comparing Alzheimer brain with appropriate controls.
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