Limits...
Epigallocatechin-3-gallate rapidly remodels PAP85-120, SEM1(45-107), and SEM2(49-107) seminal amyloid fibrils.

Castellano LM, Hammond RM, Holmes VM, Weissman D, Shorter J - Biol Open (2015)

Bottom Line: Here, we confirm that the green tea polyphenol, epigallocatechin-3-gallate (EGCG), slowly remodels fibrils formed by PAP248-286 termed SEVI (semen derived enhancer of viral infection) and also exerts a direct anti-viral effect.We elucidate for the first time that EGCG remodels PAP85-120, SEM1(45-107), and SEM2(49-107) fibrils more rapidly than SEVI fibrils.The combined anti-amyloid and anti-viral properties of EGCG could have utility in preventing HIV transmission.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Biophysics, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA Pharmacology Graduate Group, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.

No MeSH data available.


Related in: MedlinePlus

EGCG slowly remodels SEVI fibrils into non-amyloid structures. (A) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by ThT fluorescence. Values represent means±s.e.m. (n=4). (B) Transmission electron micrographs of SEVI fibrils incubated with buffer or EGCG for 2 h or 24 h. Scale bar: 2 µm. (C) SEVI fibrils (20 µM) were incubated with buffer for 6 h or 24 h or EGCG (200 µM) for 6 h or 24 h, and the resulting products were used to seed soluble PAP248-286 (1 mM, 0.1% fibril seed) fibrillization. Buffer conditions lacking fibril seed were included. Fibril assembly was monitored by ThT fluorescence. Values represent means±s.e.m. (n=4). (D-F) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by anti-amyloid (OC) immunoreactivity (D), turbidity (E), or sedimentation analysis (F). Values represent means±s.e.m. (n=3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4582112&req=5

BIO010215F1: EGCG slowly remodels SEVI fibrils into non-amyloid structures. (A) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by ThT fluorescence. Values represent means±s.e.m. (n=4). (B) Transmission electron micrographs of SEVI fibrils incubated with buffer or EGCG for 2 h or 24 h. Scale bar: 2 µm. (C) SEVI fibrils (20 µM) were incubated with buffer for 6 h or 24 h or EGCG (200 µM) for 6 h or 24 h, and the resulting products were used to seed soluble PAP248-286 (1 mM, 0.1% fibril seed) fibrillization. Buffer conditions lacking fibril seed were included. Fibril assembly was monitored by ThT fluorescence. Values represent means±s.e.m. (n=4). (D-F) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by anti-amyloid (OC) immunoreactivity (D), turbidity (E), or sedimentation analysis (F). Values represent means±s.e.m. (n=3).

Mentions: The small molecule EGCG, a potent antioxidant and polyphenol found in green tea, has previously been shown to dose-dependently disassemble SEVI fibrils over 24–48 h (Hauber et al., 2009). We confirmed this gradual disassembly, as a drastic decrease in thioflavin-T (ThT) fluorescence intensity was not observed until SEVI fibrils were treated with a ten-fold excess of EGCG for 24 h (Fig. 1A). Transmission electron microscopy (TEM) verified that fibrils were still the predominant species present after a 2 h treatment with EGCG (Fig. 1B). Furthermore, we found that SEVI fibrils pre-treated with EGCG for 6 h could still effectively ‘seed’ the fibrillization of monomeric PAP248-286 (Fig. 1C). Thus, EGCG is unable to eliminate self-templating activity or remodel SEVI into a non-amyloid form on this timescale (Fig. 1C). After a longer 24 h treatment, however, a striking change in morphology was observed by TEM, where significantly smaller oligomeric structures were observed in place of fibrils (Fig. 1B). SEVI fibrils pre-treated with EGCG for 24 h could no longer seed the assembly of PAP248-286 (Fig. 1C). We confirmed slow remodeling of SEVI fibrils by EGCG after 24 h, but not at earlier times, using three separate measures: immunoreactivity to the anti-amyloid OC antibody (Kayed et al., 2007) (Fig. 1D), turbidity (Fig. 1E), and sedimentation analysis (Fig. 1F). Thus, we confirm previous observations that EGCG remodels SEVI fibrils (Hauber et al., 2009).Fig. 1.


Epigallocatechin-3-gallate rapidly remodels PAP85-120, SEM1(45-107), and SEM2(49-107) seminal amyloid fibrils.

Castellano LM, Hammond RM, Holmes VM, Weissman D, Shorter J - Biol Open (2015)

EGCG slowly remodels SEVI fibrils into non-amyloid structures. (A) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by ThT fluorescence. Values represent means±s.e.m. (n=4). (B) Transmission electron micrographs of SEVI fibrils incubated with buffer or EGCG for 2 h or 24 h. Scale bar: 2 µm. (C) SEVI fibrils (20 µM) were incubated with buffer for 6 h or 24 h or EGCG (200 µM) for 6 h or 24 h, and the resulting products were used to seed soluble PAP248-286 (1 mM, 0.1% fibril seed) fibrillization. Buffer conditions lacking fibril seed were included. Fibril assembly was monitored by ThT fluorescence. Values represent means±s.e.m. (n=4). (D-F) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by anti-amyloid (OC) immunoreactivity (D), turbidity (E), or sedimentation analysis (F). Values represent means±s.e.m. (n=3).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4582112&req=5

BIO010215F1: EGCG slowly remodels SEVI fibrils into non-amyloid structures. (A) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by ThT fluorescence. Values represent means±s.e.m. (n=4). (B) Transmission electron micrographs of SEVI fibrils incubated with buffer or EGCG for 2 h or 24 h. Scale bar: 2 µm. (C) SEVI fibrils (20 µM) were incubated with buffer for 6 h or 24 h or EGCG (200 µM) for 6 h or 24 h, and the resulting products were used to seed soluble PAP248-286 (1 mM, 0.1% fibril seed) fibrillization. Buffer conditions lacking fibril seed were included. Fibril assembly was monitored by ThT fluorescence. Values represent means±s.e.m. (n=4). (D-F) Preformed SEVI fibrils (20 µM) were incubated with buffer or EGCG (200 µM) for 0–24 h. Fibril integrity was assessed by anti-amyloid (OC) immunoreactivity (D), turbidity (E), or sedimentation analysis (F). Values represent means±s.e.m. (n=3).
Mentions: The small molecule EGCG, a potent antioxidant and polyphenol found in green tea, has previously been shown to dose-dependently disassemble SEVI fibrils over 24–48 h (Hauber et al., 2009). We confirmed this gradual disassembly, as a drastic decrease in thioflavin-T (ThT) fluorescence intensity was not observed until SEVI fibrils were treated with a ten-fold excess of EGCG for 24 h (Fig. 1A). Transmission electron microscopy (TEM) verified that fibrils were still the predominant species present after a 2 h treatment with EGCG (Fig. 1B). Furthermore, we found that SEVI fibrils pre-treated with EGCG for 6 h could still effectively ‘seed’ the fibrillization of monomeric PAP248-286 (Fig. 1C). Thus, EGCG is unable to eliminate self-templating activity or remodel SEVI into a non-amyloid form on this timescale (Fig. 1C). After a longer 24 h treatment, however, a striking change in morphology was observed by TEM, where significantly smaller oligomeric structures were observed in place of fibrils (Fig. 1B). SEVI fibrils pre-treated with EGCG for 24 h could no longer seed the assembly of PAP248-286 (Fig. 1C). We confirmed slow remodeling of SEVI fibrils by EGCG after 24 h, but not at earlier times, using three separate measures: immunoreactivity to the anti-amyloid OC antibody (Kayed et al., 2007) (Fig. 1D), turbidity (Fig. 1E), and sedimentation analysis (Fig. 1F). Thus, we confirm previous observations that EGCG remodels SEVI fibrils (Hauber et al., 2009).Fig. 1.

Bottom Line: Here, we confirm that the green tea polyphenol, epigallocatechin-3-gallate (EGCG), slowly remodels fibrils formed by PAP248-286 termed SEVI (semen derived enhancer of viral infection) and also exerts a direct anti-viral effect.We elucidate for the first time that EGCG remodels PAP85-120, SEM1(45-107), and SEM2(49-107) fibrils more rapidly than SEVI fibrils.The combined anti-amyloid and anti-viral properties of EGCG could have utility in preventing HIV transmission.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Biophysics, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA Pharmacology Graduate Group, Perelman School of Medicine at the University of Pennsylvania, Philadelphia, PA 19104, USA.

No MeSH data available.


Related in: MedlinePlus