Molecular basis of a novel renal amyloidosis due to N184K gelsolin variant

Bonì, Francesco; Milani, Mario; Porcari, Riccardo; Barbiroli, Alberto; Ricagno, Stéfano; Rosa, Matteo de

doi:10.1038/srep33463

Cited by 12 publications

(47 citation statements)

References 30 publications

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“…The reasons behind the tissue-specificity of the GSN variants is not yet fully understood. G2 carrying the N184K substitution is still able to bind Ca 2+ , and the geometry of the binding site matches that of the wild type (WT) protein (Bonì et al 2016). Destabilization of the N184K G2 variant seems to be caused by the remodeling of the H-bond network in the core of the domain (Fig.…”

Section: Introductionmentioning

confidence: 79%

“…Thermal stability of GSN variants was evaluated as previously reported (Bonì et al 2016(Bonì et al , 2018Giorgino et al 2019). Briefly, proteins were diluted to 0.2 mg/ml in 20 mM HEPES, pH 7.4, 100 mM NaCl and either 1 mM EDTA or 1 mM CaCl2.…”

Section: Thermal Denaturation Monitored By Circular Dichroism Spectromentioning

confidence: 99%

“…1). The different mechanism of mutation-induced destabilization of G2, however, causes susceptibility to furin proteolysis comparable to that induced by D187N/Y substitutions (Bonì et al 2016), suggesting that the furin/MMPs proteolytic cascade is also responsible for the aggregation of the N184K variant.…”

Section: Introductionmentioning

confidence: 96%

“…The active/open conformation of GSN is a flexible structure, with the domains mostly connected by loose linkers (Ashish et al 2007). For this reason, many structural and functional studies relied on the use of the isolated domains (Isaacson et al 1999;Kazmirski et al 2000Kazmirski et al , 2002Ratnaswamy et al 2001;Huff et al 2003;Bonì et al 2016Bonì et al , 2018Giorgino et al 2019). In addition to Ca 2+ , other intracellular secondary messengers (Badmalia et al 2017;Szatmári et al 2018;Patel et al 2018); as well as environmental factors, such as temperature and pH, (Garg et al 2011;Badmalia et al 2017) modulate GSN localization, conformation and/or activity.…”

Section: Introductionmentioning

confidence: 99%

“…The D187N, D187Y substitutions impair Ca 2+ binding to G2, destabilize the G2 fold and increase its conformational flexibility ( Fig. 1) (Kazmirski et al 2000(Kazmirski et al , 2002Ratnaswamy et al 2001;Chen et al 2001;Huff et al 2003;Bonì et al 2016;Giorgino et al 2019). In the Golgi, high concentrations of Ca 2+ activate GSN and the mutation-induced G2 destabilization promotes its aberrant cleavage by the furin protease (Chen et al 2001).…”

Section: Introductionmentioning

confidence: 99%

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The structure of N184K amyloidogenic variant of gelsolin highlights the role of the H-bond network for protein stability and aggregation properties

et al. 2019

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Mutations in the gelsolin protein are responsible for a rare conformational disease known as AGel amyloidosis. Four of these mutations are hosted by the second domain of the protein (G2): D187N/Y, G167R and N184K. The impact of the latter has been so far evaluated only by studies on the isolated G2. Here we report the characterization of full-length gelsolin carrying the N184K mutation and compare the findings with those obtained on the wild type and the other variants.The crystallographic structure of the N184K variant in the Ca 2+ -free conformation shows remarkable similarities with the wild type protein. Only minimal local rearrangements can be observed and the mutant is as efficient as the wild type in severing filamentous actin. However, thermal stability of the pathological variant is compromised in the Ca 2+ -free conditions. These data suggest that the N to K substitution causes a local disruption of the H-bond network in the 2 core of the G2 domain. Such a subtle rearrangement of the connections does not lead to significant conformational changes but severely affects the stability of the protein.

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Section: Introductionmentioning

confidence: 79%

Section: Thermal Denaturation Monitored By Circular Dichroism Spectromentioning

confidence: 99%

Section: Introductionmentioning

confidence: 96%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The structure of N184K amyloidogenic variant of gelsolin highlights the role of the H-bond network for protein stability and aggregation properties

et al. 2019

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Aggregation of gelsolin wild-type and G167K/R, N184K, and D187N/Y mutant peptides and inhibition

et al. 2021

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Nanobody interaction unveils structure, dynamics and proteotoxicity of the Finnish-type amyloidogenic gelsolin variant

Giorgino

Mattioni

Hassan

et al. 2019

Biochimica et Biophysica Acta (BBA) - Molecular Basis of Diseas

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§ These authors contributed equally to this work. Highlights• D187N gelsolin variant is responsible for the most common form of AGel amyloidosis • We obtained the crystal structure of the second domain of D187N in complex with a nanobody• D187N substitution increases the conformational flexibility of the protein • Nanobody binding shifts D187N towards a native-like conformation• The same nanobody binds to and protects other gelsolin pathological variants (N184K and G167R)• The nanobody protects from the toxicity induced by gelsolin mutants in C. elegans AbstractAGel amyloidosis, formerly known as familial amyloidosis of the Finnish-type, is caused by pathological aggregation of proteolytic fragments of plasma gelsolin. So far, four mutations in the gelsolin gene have been reported as responsible for the disease. Although D187N is the first identified variant and the best characterized, its structure has been hitherto elusive. Exploiting a recently-developed nanobody targeting gelsolin, we were able to stabilize the G2 domain of the D187N protein and obtained, for the first time, its high-resolution crystal structure. In the nanobody-stabilized conformation, the main effect of the D187N substitution is the impairment of the calcium binding capability, leading to a destabilization of the C-terminal tail of G2. However, molecular dynamics simulations show that in the absence of the nanobody, D187N-mutated G2 further misfolds, ultimately exposing its hydrophobic core and the furin cleavage site. The nanobody's protective effect is based on the enhancement of the thermodynamic stability of different G2 mutants (D187N, G167R and N184K). In particular, the nanobody reduces the flexibility of dynamic stretches, and most notably decreases the conformational entropy of the Cterminal tail, otherwise stabilized by the presence of the Ca 2+ ion. A Caenorhabditis elegans-based assay was also applied to quantify the proteotoxic potential of the mutants and determine whether nanobody stabilization translates into a biologically relevant effect. Successful protection from G2 toxicity in vivo points to the use of C. elegans as a tool for investigating the mechanisms underlying AGel amyloidosis and rapidly screen new therapeutics.

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Molecular basis of a novel renal amyloidosis due to N184K gelsolin variant

Cited by 12 publications

References 30 publications

The structure of N184K amyloidogenic variant of gelsolin highlights the role of the H-bond network for protein stability and aggregation properties

The structure of N184K amyloidogenic variant of gelsolin highlights the role of the H-bond network for protein stability and aggregation properties

Aggregation of gelsolin wild-type and G167K/R, N184K, and D187N/Y mutant peptides and inhibition

Nanobody interaction unveils structure, dynamics and proteotoxicity of the Finnish-type amyloidogenic gelsolin variant

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