Amyloidogenicity of beta A4 and beta A4-bearing amyloid protein precursor fragments by metal-catalyzed oxidation.
Previously we have shown that the COOH-terminal 100 residues (A4CT) of the amyloid protein precursor (APP), which carry the sequence of the amyloid beta A4 protein of Alzheimer's disease at N-terminal position, form highly insoluble aggregates if expressed in the rabbit reticulocyte lysate and analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (Dyrks, T., Weidemann, A., Multhaup, G., Salbaum, J.M., Lemaire, H.-G., Kang, J., Müller-Hill, B., Masters, C. L., and Beyreuther, K. (1988) EMBO J. 7, 949-957). Here we report that aggregation of this COOH-terminal APP fragment A4CT and also of beta A4 itself depends on additional factors. In contrast to the reticulocyte expression system, expression of A4CT and beta A4 in the wheat germ expression system resulted in only monomeric forms. We have identified the factors which are capable of transforming both soluble A4CT and beta A4 into insoluble and aggregating molecules. Monomeric A4CT or beta A4 expressed in the wheat germ lysate could be transformed into aggregating molecules by the addition of metal-catalyzed oxidation systems. The addition of radical scavengers such as ascorbic acid, trolox, and amino acids prevented the aggregation process induced by the radical initiators. Thus, the aggregation of amyloidogenic APP fragments if analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis requires amino acid oxidation and protein cross-linking induced by radical generation systems.
Dyrks
T
T
Center for Molecular Biology, University of Heidelberg, Germany.
Dyrks
E
E
Hartmann
T
T
Masters
C
C
Beyreuther
K
K
D016564:Amyloid beta-Protein Precursor / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D000818:Animals; D004591:Electrophoresis, Polyacrylamide Gel; D005822:Genetic Vectors; D006418:Heme / Q000494:pharmacology; D006454:Hemoglobins / Q000494:pharmacology; D006801:Humans; D007501:Iron / Q000494:pharmacology*; D010084:Oxidation-Reduction; D010446:Peptide Fragments / Q000302:isolation & purification / Q000378:metabolism; D010957:Plasmids; D014176:Protein Biosynthesis; D011817:Rabbits; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D012156:Reticulocytes / Q000378:metabolism; D014158:Transcription, Genetic; D014908:Triticum
D016564:Amyloid beta-Protein Precursor; D006454:Hemoglobins; D010446:Peptide Fragments; D011994:Recombinant Proteins; D006418:Heme; D007501:Iron