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Alan R Fersht - One of the best experts on this subject based on the ideXlab platform.
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loss of a metal binding site in gelsolin leads to Familial Amyloidosis finnish type
Nature Structural & Molecular Biology, 2002Co-Authors: Steven L Kazmirski, Rivka L Isaacson, Ashley M Buckle, Christopher M Johnson, Valerie Daggett, Alan R FershtAbstract:Mutations in domain 2 (D2, residues 151-266) of the actin-binding protein gelsolin cause Familial Amyloidosis-Finnish type (FAF). These mutations, D187N or D187Y, lead to abnormal proteolysis of plasma gelsolin at residues 172-173 and a second hydrolysis at residue 243, resulting in an amyloidogenic fragment. Here we present the structure of human gelsolin D2 at 1.65 A and find that Asp 187 is part of a Cd2+ metal-binding site. Two Ca2+ ions are required for a conformational transition of gelsolin to its active form. Differential scanning calorimetry (DSC) and molecular dynamics (MD) simulations suggest that the Cd2+-binding site in D2 is one of these two Ca2+-binding sites and is essential to the stability of D2. Mutation of Asp 187 to Asn disrupts Ca2+ binding in D2, leading to instabilities upon Ca2+ activation. These instabilities make the domain a target for aberrant proteolysis, thereby enacting the first step in the cascade leading to FAF.
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elucidating the mechanism of Familial Amyloidosis finnish type nmr studies of human gelsolin domain 2
Proceedings of the National Academy of Sciences of the United States of America, 2000Co-Authors: Steven L Kazmirski, Rivka L Isaacson, Mark J Howard, Alan R FershtAbstract:Familial Amyloidosis-Finnish type (FAF) results from a single mutation at residue 187 (D187N or D187Y) within domain 2 of the actin-regulating protein gelsolin. The mutation somehow allows a masked cleavage site to be exposed, leading to the first step in the formation of an amyloidogenic fragment. We have performed NMR experiments investigating structural and dynamic changes between wild-type (WT) and D187N gelsolin domain 2 (D2). On mutation, no significant structural or dynamic changes occur at or near the cleavage site. Areas in conformational exchange are observed between beta-strand 4 and alpha-helix 1 and within the loop region following beta-strand 5. Chemical shift differences are noted along the face of alpha-helix 1 that packs onto the beta-sheet, suggesting an altered conformation. Conformational changes within these areas can have an effect on actin binding and may explain why D187N gelsolin is inactive. [(1)H-(15)N] nuclear Overhauser effect and chemical shift data suggest that the C-terminal tail of D187N gelsolin D2 is less structured than WT by up to six residues. In the crystal structure of equine gelsolin, the C-terminal tail of D2 lies across a large cleft between domains 1 and 2 where the masked cleavage site sits. We propose that the D187N mutation destabilizes the C-terminal tail of D2 resulting in a more exposed cleavage site leading to the first proteolysis step in the formation of the amyloidogenic fragment.
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equilibria and kinetics of folding of gelsolin domain 2 and mutants involved in Familial Amyloidosis finnish type
Proceedings of the National Academy of Sciences of the United States of America, 1999Co-Authors: Rivka L Isaacson, Alan G Weeds, Alan R FershtAbstract:Mutations D187N and D187Y in domain 2 of the actin-regulating protein gelsolin cause Familial Amyloidosis–Finnish type (FAF). We have constructed and expressed a recombinant version of gelsolin domain 2 that is sufficiently stable for kinetic and equilibrium measurements. The wild-type domain and the two amyloidogenic mutants fold via simple two-state kinetics without the accumulation of an intermediate. Unfolding kinetics exhibits significant curvature with increasing urea concentration, indicating that the transition state for unfolding becomes more native-like under increasingly denaturing conditions in accordance with the Hammond postulate. Mutations D187N and D187Y destabilize gelsolin domain 2 by 1.22 and 2.16 kcal⋅mol−1 (1 kcal = 4.18 kJ) respectively. The mutations do not prevent disulfide bond formation despite their direct contiguity with a cysteine residue involved in disulfide linkage. The destabilization conferred on gelsolin domain 2 by the FAF mutations is sufficient to predict that an appreciable fraction is unfolded and, therefore, extremely susceptible to proteolysis at body temperature.
Paulo Torres - One of the best experts on this subject based on the ideXlab platform.
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ophthalmological manifestations in hereditary transthyretin attr v30m carriers a review of 513 cases
Amyloid, 2015Co-Authors: Joao Melo Beirao, Paulo Costa, Jorge Malheiro, Carolina Lemos, Idalina Beirao, Paulo TorresAbstract:AbstractPurpose: Assessment of ocular involvement in transthyretin-related Familial Amyloidosis with polyneuropathy (FAP) in a large cohort of Portuguese patients.Methods: We reviewed the medical records of 513 Portuguese FAP mutation carriers, at the Ophthalmology Service, Centro Hospitalar do Porto, between 1 January 2008 and 31 January 2013. Abnormal conjunctiva vessels (ACV), Schirmer test, tear break-up time (TBUT), amyloid deposition on the iris (DAI), scalloped iris, amyloid deposition on the anterior capsule of the lens (DAL), vitreous Amyloidosis, retinal amyloid angiopathy and glaucoma were evaluated and registered.Results: Of the 513 carriers, 477 (93%) had clinical systemic disease with a median duration of 9.3 (5.1–13.7) years and 247 were men. Of these, 343 (72%) had been liver transplanted, on median of 6.6 (3.3–10.8) years before inclusion in this study. No ocular abnormalities were identified in the asymptomatic carriers (7%). The abnormalities observed with decreasing frequency were abno...
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Vitreous surgery impact in glaucoma development in liver transplanted Familial Amyloidosis ATTR V30M Portuguese patients.
Amyloid : the international journal of experimental and clinical investigation : the official journal of the International Society of Amyloidosis, 2012Co-Authors: Nuno M. Beirão, Idalina Beirâo, Paulo Costa, Maria E. Matos, Maria João Menéres, Paulo TorresAbstract:Purpose: Familial Amyloidosis with polyneuropathy (FAP) sometimes courses with vitreous amyloid. The aim of this study was to evaluate the incidence of glaucoma after vitrectomy in FAP patients. Methods: A total of 79 eyes of 42 liver transplanted FAP patients and 16 eyes of 16 non-FAP patients with rhegmatogenous retina detachment were collected. The patients were divided in to three groups: Group I – FAP patients with vitreous opacities submitted to vitrectomy, Group II – FAP patients without vitreous opacities and not submitted to vitrectomy and, Group III – non-FAP patients with rhegmatogenous retinal detachment submitted to vitrectomy. The Group I was subdivided into: Ia – “complete” vitrectomy; Ib – “incomplete” vitrectomy. The onset of glaucoma was considered when the intraocular pressure level was higher than 21 mmHg, with concomitant visual field abnormalities and optic nerve cupping. Results: Post vitrectomy glaucoma was more frequent in Group I (56.1%) than in Group III (12.5%) and in Group II ...
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anticipation of presbyopia in portuguese Familial Amyloidosis attr v30m
Amyloid, 2011Co-Authors: Melo Beirao, Eduarda Matos, Idalina Beirâo, Paulo Costa, Paulo TorresAbstract:The aim of this study was to evaluate if Portuguese patients with Familial Amyloidosis, liver transplanted and not, have an earlier development of presbyopia compared with a normal population and its relation with the presence or the absence of anterior capsule opacification of the lens. This study was performed to evaluate if Portuguese patients with Familial Amyloidosis and in a blood donors population (control group). Three hundred and fifty-six subjects, 144 amyloidotic patients and 212 healthy individuals, were evaluated for the need of plus lenses for normal near reading (Jaeger chart 1 at 33 cm). In Familial Amyloidosis patients, the value of the add-power was related to age, liver transplantation status, and presence of visible anterior capsule opacification of the lens. In both groups, the value of add-power was positively correlated with age (r = 0.91; P < 0.005). Familial Amyloidosis patients require more add-power than control individuals of similar age, and need to use reading glasses at earl...
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recurrence of vitreous Amyloidosis and need of surgical reintervention in portuguese patients with Familial Amyloidosis attr v30m
Retina-the Journal of Retinal and Vitreous Diseases, 2011Co-Authors: Nuno M. Beirão, Eduarda Matos, Idalina Beirâo, Paulo Costa, Paulo TorresAbstract:PURPOSE Vitreous amyloid deposits are one of the most common ocular manifestations of Familial Amyloidosis ATTR V30M (FAP-I), which can be the only manifestation of the disease and can appear even after liver transplantation. Removal by vitrectomy is usually performed, but vitreous amyloid recurrence has been frequently reported. This study was undertaken to evaluate the recurrence of vitreous Amyloidosis and its relationship with the degree of previous vitreous removal. METHODS Fifty-four vitrectomized eyes from 32 patients with FAP-I were evaluated in the course of a follow-up period of 30.7 ± 17.2 months (range, 8-78; median = 30 months). An extensive, as possible, vitrectomy with indentation was performed in 41 eyes (complete), and in the others 13 eyes only a vitrectomy without indentation (incomplete) was performed. The parameters evaluated were the incidence of amyloid deposits and visual outcomes. RESULTS A noteworthy visual acuity gain was observed, although a few patients had a subsequent decrease of visual acuity related to new vitreous amyloid deposition in the visual axis. These new amyloid deposits did not occur in eyes that had undergone extensive vitreous removal, but only in nonextensive vitrectomized eyes (P < 0.001). CONCLUSION Recurrence of amyloid deposition only occurred in nonextensive vitrectomized eyes and represents a false recurrence associated with incomplete vitrectomy.
Rivka L Isaacson - One of the best experts on this subject based on the ideXlab platform.
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loss of a metal binding site in gelsolin leads to Familial Amyloidosis finnish type
Nature Structural & Molecular Biology, 2002Co-Authors: Steven L Kazmirski, Rivka L Isaacson, Ashley M Buckle, Christopher M Johnson, Valerie Daggett, Alan R FershtAbstract:Mutations in domain 2 (D2, residues 151-266) of the actin-binding protein gelsolin cause Familial Amyloidosis-Finnish type (FAF). These mutations, D187N or D187Y, lead to abnormal proteolysis of plasma gelsolin at residues 172-173 and a second hydrolysis at residue 243, resulting in an amyloidogenic fragment. Here we present the structure of human gelsolin D2 at 1.65 A and find that Asp 187 is part of a Cd2+ metal-binding site. Two Ca2+ ions are required for a conformational transition of gelsolin to its active form. Differential scanning calorimetry (DSC) and molecular dynamics (MD) simulations suggest that the Cd2+-binding site in D2 is one of these two Ca2+-binding sites and is essential to the stability of D2. Mutation of Asp 187 to Asn disrupts Ca2+ binding in D2, leading to instabilities upon Ca2+ activation. These instabilities make the domain a target for aberrant proteolysis, thereby enacting the first step in the cascade leading to FAF.
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elucidating the mechanism of Familial Amyloidosis finnish type nmr studies of human gelsolin domain 2
Proceedings of the National Academy of Sciences of the United States of America, 2000Co-Authors: Steven L Kazmirski, Rivka L Isaacson, Mark J Howard, Alan R FershtAbstract:Familial Amyloidosis-Finnish type (FAF) results from a single mutation at residue 187 (D187N or D187Y) within domain 2 of the actin-regulating protein gelsolin. The mutation somehow allows a masked cleavage site to be exposed, leading to the first step in the formation of an amyloidogenic fragment. We have performed NMR experiments investigating structural and dynamic changes between wild-type (WT) and D187N gelsolin domain 2 (D2). On mutation, no significant structural or dynamic changes occur at or near the cleavage site. Areas in conformational exchange are observed between beta-strand 4 and alpha-helix 1 and within the loop region following beta-strand 5. Chemical shift differences are noted along the face of alpha-helix 1 that packs onto the beta-sheet, suggesting an altered conformation. Conformational changes within these areas can have an effect on actin binding and may explain why D187N gelsolin is inactive. [(1)H-(15)N] nuclear Overhauser effect and chemical shift data suggest that the C-terminal tail of D187N gelsolin D2 is less structured than WT by up to six residues. In the crystal structure of equine gelsolin, the C-terminal tail of D2 lies across a large cleft between domains 1 and 2 where the masked cleavage site sits. We propose that the D187N mutation destabilizes the C-terminal tail of D2 resulting in a more exposed cleavage site leading to the first proteolysis step in the formation of the amyloidogenic fragment.
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equilibria and kinetics of folding of gelsolin domain 2 and mutants involved in Familial Amyloidosis finnish type
Proceedings of the National Academy of Sciences of the United States of America, 1999Co-Authors: Rivka L Isaacson, Alan G Weeds, Alan R FershtAbstract:Mutations D187N and D187Y in domain 2 of the actin-regulating protein gelsolin cause Familial Amyloidosis–Finnish type (FAF). We have constructed and expressed a recombinant version of gelsolin domain 2 that is sufficiently stable for kinetic and equilibrium measurements. The wild-type domain and the two amyloidogenic mutants fold via simple two-state kinetics without the accumulation of an intermediate. Unfolding kinetics exhibits significant curvature with increasing urea concentration, indicating that the transition state for unfolding becomes more native-like under increasingly denaturing conditions in accordance with the Hammond postulate. Mutations D187N and D187Y destabilize gelsolin domain 2 by 1.22 and 2.16 kcal⋅mol−1 (1 kcal = 4.18 kJ) respectively. The mutations do not prevent disulfide bond formation despite their direct contiguity with a cysteine residue involved in disulfide linkage. The destabilization conferred on gelsolin domain 2 by the FAF mutations is sufficient to predict that an appreciable fraction is unfolded and, therefore, extremely susceptible to proteolysis at body temperature.
Robert Robinson - One of the best experts on this subject based on the ideXlab platform.
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the role of gelsolin domain 3 in Familial Amyloidosis finnish type
Proceedings of the National Academy of Sciences of the United States of America, 2019Co-Authors: Jan Gettemans, Habiba Zorgati, Marten Larsson, Weitong Ren, Adelene Y L Sim, Jonathan M Grimes, Robert RobinsonAbstract:In the disease Familial Amyloidosis, Finnish type (FAF), also known as AGel Amyloidosis (AGel), the mechanism by which point mutations in the calcium-regulated actin-severing protein gelsolin lead to furin cleavage is not understood in the intact protein. Here, we provide a structural and biochemical characterization of the FAF variants. X-ray crystallography structures of the FAF mutant gelsolins demonstrate that the mutations do not significantly disrupt the calcium-free conformations of gelsolin. Small-angle X-ray–scattering (SAXS) studies indicate that the FAF calcium-binding site mutants are slower to activate, whereas G167R is as efficient as the wild type. Actin-regulating studies of the gelsolins at the furin cleavage pH (6.5) show that the mutant gelsolins are functional, suggesting that they also adopt relatively normal active conformations. Deletion of gelsolin domains leads to sensitization to furin cleavage, and nanobody-binding protects against furin cleavage. These data indicate instability in the second domain of gelsolin (G2), since loss or gain of G2-stabilizing interactions impacts the efficiency of cleavage by furin. To demonstrate this principle, we engineered non-FAF mutations in G3 that disrupt the G2-G3 interface in the calcium-activated structure. These mutants led to increased furin cleavage. We carried out molecular dynamics (MD) simulations on the FAF and non-FAF mutant G2-G3 fragments of gelsolin. All mutants showed an increase in the distance between the center of masses of the 2 domains (G2 and G3). Since G3 covers the furin cleavage site on G2 in calcium-activated gelsolin, this suggests that destabilization of this interface is a critical step in cleavage.
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the crystal structure of plasma gelsolin implications for actin severing capping and nucleation
Cell, 1997Co-Authors: Leslie D Burtnick, Edward K Koepf, J M Grimes, E Y Jones, D I Stuart, P J Mclaughlin, Robert RobinsonAbstract:The structure of gelsolin has been determined by crystallography and comprises six structurally related domains that, in a Ca2+-free environment, pack together to form a compact globular structure in which the putative actin-binding sequences are not sufficiently exposed to enable binding to occur. We propose that binding Ca2+ can release the connections that join the N- and C-terminal halves of gelsolin, enabling each half to bind actin relatively independently. Domain shifts are proposed in response to Ca2+ as bases for models of how gelsolin acts to sever, cap, or nucleate F-actin filaments. The structure also invites discussion of polyphosphoinositide binding to segment 2 and suggests how mutation at Asp-187 could initiate a series of events that lead to deposition of amyloid plaques, as observed in victims of Familial Amyloidosis (Finnish type).
Paulo Costa - One of the best experts on this subject based on the ideXlab platform.
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ophthalmological manifestations in hereditary transthyretin attr v30m carriers a review of 513 cases
Amyloid, 2015Co-Authors: Joao Melo Beirao, Paulo Costa, Jorge Malheiro, Carolina Lemos, Idalina Beirao, Paulo TorresAbstract:AbstractPurpose: Assessment of ocular involvement in transthyretin-related Familial Amyloidosis with polyneuropathy (FAP) in a large cohort of Portuguese patients.Methods: We reviewed the medical records of 513 Portuguese FAP mutation carriers, at the Ophthalmology Service, Centro Hospitalar do Porto, between 1 January 2008 and 31 January 2013. Abnormal conjunctiva vessels (ACV), Schirmer test, tear break-up time (TBUT), amyloid deposition on the iris (DAI), scalloped iris, amyloid deposition on the anterior capsule of the lens (DAL), vitreous Amyloidosis, retinal amyloid angiopathy and glaucoma were evaluated and registered.Results: Of the 513 carriers, 477 (93%) had clinical systemic disease with a median duration of 9.3 (5.1–13.7) years and 247 were men. Of these, 343 (72%) had been liver transplanted, on median of 6.6 (3.3–10.8) years before inclusion in this study. No ocular abnormalities were identified in the asymptomatic carriers (7%). The abnormalities observed with decreasing frequency were abno...
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Vitreous surgery impact in glaucoma development in liver transplanted Familial Amyloidosis ATTR V30M Portuguese patients.
Amyloid : the international journal of experimental and clinical investigation : the official journal of the International Society of Amyloidosis, 2012Co-Authors: Nuno M. Beirão, Idalina Beirâo, Paulo Costa, Maria E. Matos, Maria João Menéres, Paulo TorresAbstract:Purpose: Familial Amyloidosis with polyneuropathy (FAP) sometimes courses with vitreous amyloid. The aim of this study was to evaluate the incidence of glaucoma after vitrectomy in FAP patients. Methods: A total of 79 eyes of 42 liver transplanted FAP patients and 16 eyes of 16 non-FAP patients with rhegmatogenous retina detachment were collected. The patients were divided in to three groups: Group I – FAP patients with vitreous opacities submitted to vitrectomy, Group II – FAP patients without vitreous opacities and not submitted to vitrectomy and, Group III – non-FAP patients with rhegmatogenous retinal detachment submitted to vitrectomy. The Group I was subdivided into: Ia – “complete” vitrectomy; Ib – “incomplete” vitrectomy. The onset of glaucoma was considered when the intraocular pressure level was higher than 21 mmHg, with concomitant visual field abnormalities and optic nerve cupping. Results: Post vitrectomy glaucoma was more frequent in Group I (56.1%) than in Group III (12.5%) and in Group II ...
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anticipation of presbyopia in portuguese Familial Amyloidosis attr v30m
Amyloid, 2011Co-Authors: Melo Beirao, Eduarda Matos, Idalina Beirâo, Paulo Costa, Paulo TorresAbstract:The aim of this study was to evaluate if Portuguese patients with Familial Amyloidosis, liver transplanted and not, have an earlier development of presbyopia compared with a normal population and its relation with the presence or the absence of anterior capsule opacification of the lens. This study was performed to evaluate if Portuguese patients with Familial Amyloidosis and in a blood donors population (control group). Three hundred and fifty-six subjects, 144 amyloidotic patients and 212 healthy individuals, were evaluated for the need of plus lenses for normal near reading (Jaeger chart 1 at 33 cm). In Familial Amyloidosis patients, the value of the add-power was related to age, liver transplantation status, and presence of visible anterior capsule opacification of the lens. In both groups, the value of add-power was positively correlated with age (r = 0.91; P < 0.005). Familial Amyloidosis patients require more add-power than control individuals of similar age, and need to use reading glasses at earl...
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recurrence of vitreous Amyloidosis and need of surgical reintervention in portuguese patients with Familial Amyloidosis attr v30m
Retina-the Journal of Retinal and Vitreous Diseases, 2011Co-Authors: Nuno M. Beirão, Eduarda Matos, Idalina Beirâo, Paulo Costa, Paulo TorresAbstract:PURPOSE Vitreous amyloid deposits are one of the most common ocular manifestations of Familial Amyloidosis ATTR V30M (FAP-I), which can be the only manifestation of the disease and can appear even after liver transplantation. Removal by vitrectomy is usually performed, but vitreous amyloid recurrence has been frequently reported. This study was undertaken to evaluate the recurrence of vitreous Amyloidosis and its relationship with the degree of previous vitreous removal. METHODS Fifty-four vitrectomized eyes from 32 patients with FAP-I were evaluated in the course of a follow-up period of 30.7 ± 17.2 months (range, 8-78; median = 30 months). An extensive, as possible, vitrectomy with indentation was performed in 41 eyes (complete), and in the others 13 eyes only a vitrectomy without indentation (incomplete) was performed. The parameters evaluated were the incidence of amyloid deposits and visual outcomes. RESULTS A noteworthy visual acuity gain was observed, although a few patients had a subsequent decrease of visual acuity related to new vitreous amyloid deposition in the visual axis. These new amyloid deposits did not occur in eyes that had undergone extensive vitreous removal, but only in nonextensive vitrectomized eyes (P < 0.001). CONCLUSION Recurrence of amyloid deposition only occurred in nonextensive vitrectomized eyes and represents a false recurrence associated with incomplete vitrectomy.
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kidney and anemia in Familial Amyloidosis type i
Kidney International, 2004Co-Authors: Idalina Beirâo, Paulo Costa, L Lobato, Isabel Fonseca, Paula Mendes, Manuela Silva, Fernanda Bravo, Antonio Cabrita, Graca PortoAbstract:Kidney and anemia in Familial Amyloidosis type I. Background Familial amyloid polyneuropathy (FAP) type I is caused by a mutated transthyretin (TTR V30M) and characterized by a sensorimotor and autonomic neuropathy. Renal, cardiac, and ocular abnormalities can also occur. Anemia has been described in previous reports, but its prevalence in Portuguese FAP patients is not precisely known. The aim of this study was to estimate the prevalence of anemia in FAP type I Portuguese patients and to evaluate the contribution of erythropoietin (Epo) to its genesis. Methods A retrospective cross-sectional study was undertaken to determinate the prevalence and characteristics of anemia in 165 FAP patients. For comparison analysis, 3 control groups were also evaluated, 1 group of 46 apparently healthy subjects, 1 group of 17 asymptomatic carriers of FAP-trait, and a group of 14 non-FAP patients with chronic renal insufficiency. Serum Epo levels were analyzed in all groups. Results Anemia was present in 24.8% of symptomatic FAP patients. Iron stores, B12 vitamin, and serum folate levels were normal. FAP patients presented significantly lower serum Epo levels than healthy controls ( P = 0.003). Epo levels were found lower than expected for the degree of anemia and in 17.5% were undetectable. Low Epo values were observed independently of the presence of renal failure or anemia, and sometimes preceded clinical disease. Conclusion Anemia in FAP type I is a common manifestation. The results clearly suggest a defective endogenous Epo production in the genesis of the anemia.