Active Pre-formed Fibrils for Neurodegeneration Research

Beta synuclein, like alpha synuclein, is found in presynaptic nerve terminals. It is believed to play a role in maintaining vesicular membrane curvature¹ and does not aggregate under physiological conditions.² WT beta synuclein inhibits the aggregation of alpha synuclein³ and mutant (V70M and P123H) beta synuclein has been associated with Dementia with Lewy Bodies (DLB).⁴ Human beta synuclein PFFs do not induce endogenous alpha synuclein aggregation in primary rat hippocampal neurons, meaning they could be useful as a control in alpha synuclein experiments.

Gamma synuclein is associated with neurodegenerative diseases as well as certain cancers. Gamma synuclein can seed alpha synuclein aggregation,1 and its oxidation and aggregation is seen in Dementia with Lewy bodies,2 Parkinson’s Disease,2 and ALS.3

StressMarq is in the process of developing new types of fibrils for neurodegenerative disease research as well as thoroughly characterizing our current products. We currently have several types of gamma synuclein PFFs available. This table summarizes the differences between them:

TEM of Active Human Recombinant Gamma Synuclein Protein Preformed Fibrils (Type 1) (SPR-459)

1. Surguchov A: Synucleins: are they two-edged swords? J Neurosci Res 2013, 91(2):16
2. Galvin, J.E. Axon pathology in Parkinson’s disease and Lewy body dementia hippocampus contains a-, b-, and g-synuclein. Proc Natl Acad Sci USA. 1999 Nov 9; 96(23): 13450–13455.
3. Peters OM, Millership S, Shelkovnikova TA, et al. Selective pattern of motor system damage in gamma-synuclein transgenic mice mirrors the respective pathology in amyotrophic lateral sclerosis. Neurobiol Dis 2012;48: 124–131.

Cu/Zn superoxide dismutase 1 (SOD1) misfolding and aggregation into neurotoxic species is implicated in Amyotrophic Lateral Sclerosis (ALS). Superoxide dismutase (SOD) is an endogenously produced intracellular enzyme present in almost every cell in the body (3). It works by catalyzing the dismutation of the superoxide radical O2ˉ to O2 and H2O2, which are then metabolized to H2O and O2 by catalase and glutathione peroxidase (2,5). In general, SODs play a major role in antioxidant defense mechanisms (4). There are two main types of SOD in mammalian cells. One form (SOD1) contains Cu and Zn ions as a homodimer and exists in the cytoplasm. The two subunits of 16 kDa each are linked by two cysteines forming an intra-subunit disulphide bridge (3). SOD1 misfolding and aggregation into neurotoxic species is implicated in Amyotrophic Lateral Sclerosis (ALS).

1. Adachi T., et al. (1992). Clin. Chim. Acta. 212: 89-102.
2. Barrister J.V., et al. (1987). Crit. Rev. Biochem. 22:111-180.
3. Furukawa Y., O’Halloran T. (2006). Antioxidants & Redo Signaling. Vol 8, No 5,6.
4. Gao B., et al. (2003). Am J Physiol Lung Cell Mol Physiol 284: L917-L925.
5. Hassan H.M. (1988). Free Radical Biol. Med. 5: 377-385.

StressMarq’s Active Tau Proteins

StressMarq is excited to announce the launch of new active tau proteins to help researchers study tau aggregation, a hallmark of neurodegenerative diseases including Alzheimer’s. StressMarq is the first to offer active tau preformed fibrils (PFFs) and filaments for neuroscience research.  The process of tau aggregation can be seeded by active tau PFFs, which recruit monomers to form larger tau fibrils.  This has been demonstrated in thioflavin T assays where an increase in fluorescence, indicative of tau fibrillization, is seen when active tau PFFs are combined with active tau monomers.  Certain tau PFFs have been injected into P301L mice, where they seed tau aggregation and induce tau pathology in the hippocampus.

Immunohistochemistry analysis of P301L mouse hippocampus injected with K18 P301L tau PFFs (SPR-330) shows seeding of tau pathology at injection site. AT8 (pSer202/pThr205) tau antibody shows tangle-like inclusions. Experiments performed at reMYND N.V.

Monomers and fibrils are available both in the full-length isoform of the tau protein (2N4R or Tau-441) or a truncated form (K18).  Tau-441 has a molecular weight of approximately 46 kDa, whereas K18 tau has a molecular weight of approximately 15 kDa.  dGAE is a fragment of the tau protein consisting of amino acids 297-391. It is one of the core PHF subunits,¹ includes both microtubule-binding domains and proline-rich regions, and assembles into PHF-like fibrils in vitro without additives or templates.² 

Proteins are available as wild-type or with a variety of mutations. P301S and P301L mutations occur in exon 10 and are associated with frontotemporal dementia.  The P301S mutation reduces tau’s ability to assemble microtubules, and the P301L mutation promotes beta-sheet formation and the formation of PHFs.  Both P301S and P301L mutant transgenic mouse models are used in tau research.  The K280 deletion mutation is also associated with frontotemporal dementia and promotes fibrillization into paired helical filaments (PHFs) in the absence of heparin and other inducers. The C322A mutation also increases tau’s ability to form PHFs.²

PFFs induce tau aggregation; full-length PFFs may be more effective in seeding fibrillization, but a combination of both can be particularly toxic to neurons. Most tau varieties are fibrillized using a heparin scaffold; soluble tau filaments are fibrillized using a linear anionic scaffold. dGAE tau and K18 K280 deletion tau both fibrillize without scaffolds.

1. Novak, M. et al. (1993) EMBO J. 12, 365-370.
2. Al-Hilaly, Y.K. et al. (2017) J. Mol. Biol. 429(23):3650-3665.

StressMarq is in the process of developing new types of fibrils for neurodegenerative disease research as well as thoroughly characterizing our current products. We currently have several types of tau PFFs and filaments available. This table summarizes the differences between them:

Transthyretin (TTR) misfolding is associated with amyloid diseases. Both the L55P and Y78F mutants are amyloidogenic.^1,2 L55P and Y78F TTR filaments and monomers are available. TTR aggregation can be measured via turbidity.³­

StressMarq is in the process of developing new types of fibrils for neurodegenerative disease research as well as thoroughly characterizing our current products. We currently have several types of TTR filaments available. This table summarizes the differences between them: