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The Peptide KLVFF-K₆ Promotes -Amyloid(1–40) Protofibril Growth by Association but Does Not Alter Protofibril Effects on Cellular Reduction of 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium Bromide (MTT)

Department of Neurosciences, Mayo Clinic, Jacksonville, Florida (M.A.M., M.R.C., D.K.R., T.L.R.); and Departments of Pathology and Physiology, Johns Hopkins University School of Medicine, Baltimore, Maryland (J.H.H.)

The peptide KLVFF-K₆ was observed by Lowe et al. (Biochemistry 40:7882–7889, 2001). to simultaneously enhance amyloid -protein (A) fibrillogenesis and decrease cellular toxicity, as measured in a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reduction assay. It was postulated that accelerated A aggregation and precipitation induced by KLVFF-K₆ may lead to an increase in less toxic insoluble fibrils at the expense of more toxic soluble protofibrils. In a previous study, we distinguished between two modes of protofibril growth: elongation by monomer deposition and direct protofibril-protofibril association. These growth mechanisms could be resolved by varying A monomer and NaCl concentrations. Using assays designed to isolate these distinct modes of protofibril growth, we report here that larger A aggregates formed in the presence of KLVFF-K₆ resulted from enhanced protofibril association. ³H-Radiomethylated KLVFF-K₆ bound to associated protofibrils with an apparent K_d of 180 nM, and concentrations of free [³H]KLVFF-K₆ in this range were sufficient to convert soluble protofibrils to sedimentable fibrils. However, promotion of A protofibril association by KLVFF-K₆ had no effect on A-induced decreases in cellular MTT reduction. Therefore, our data do not support the proposal that insoluble fibrils formed with KLVFF-K₆ are less toxic than soluble protofibrils. KLVFF-K₆ did not alter rates of protofibril elongation by monomer deposition. In contrast, when added to A monomers isolated with the use of size-exclusion chromatography, KLVFF-K₆ inhibited fibrillogenesis, as measured by thioflavin T fluorescence, and this inhibition was paralleled by a failure to alter cellular MTT reduction.

Address correspondence to: Terrone L. Rosenberry, Department of Neurosciences, Mayo Clinic, 4500 San Pablo Road, Jacksonville, FL 32224. E-mail: rosenberry{at}mayo.edu

This article has been cited by other articles:

				M. A. Moss, N. H. Varvel, M. R. Nichols, D. K. Reed, and T. L. Rosenberry Nordihydroguaiaretic Acid Does Not Disaggregate {beta}-Amyloid(1-40) Protofibrils but Does Inhibit Growth Arising from Direct Protofibril Association Mol. Pharmacol., September 1, 2004; 66(3): 592 - 600. [Abstract] [Full Text] [PDF]

				J. R. Kim and R. M. Murphy Mechanism of Accelerated Assembly of {beta}-Amyloid Filaments into Fibrils by KLVFFK6 Biophys. J., May 1, 2004; 86(5): 3194 - 3203. [Abstract] [Full Text] [PDF]