New findings on GFP-like protein application as fluorescent tags: Fibrillogenesis, oligomerization, and amorphous aggregation

Sulatsky, Maksim I.; Михайлова, Е. В.; Kuznetsova, Irina М.; Turoverov, Konstantin K.; Stepanenko, Olga V.; Sulatskaya, Anna I.

doi:10.1016/j.ijbiomac.2021.10.107

Cited by 16 publications

(14 citation statements)

References 44 publications

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“…The formation of amyloid fibrils in vitro under the influence of external factors (temperature and low acidity) was also shown for the superfolder GFP (sfGFP) with a native β-barrel structure [ 187 ]. The unfolding/refolding of GFP-like proteins induced by various chemical denaturants and heating is complicated by the formation of several intermediate partially folded states [ 188 , 189 , 190 , 191 , 192 , 193 , 194 ].…”

Section: The Transition Of β-Barrel Proteins To Amyloidsmentioning

confidence: 99%

“…The exposure of clusters of hydrophobic amino acids of GFP in an intermediate state is considered to be the main reason for abnormal protein aggregation upon heating [ 197 ]. Interestingly, amyloids formed from sfGFP in vitro lose their green fluorescence, and are toxic to mammalian cells [ 187 ]. It is unclear whether GFP-like proteins can form bona fide amyloid fibrils in vivo but, as was mentioned above, they show high oligomerization and, in several cases, aggregation propensity.…”

Section: The Transition Of β-Barrel Proteins To Amyloidsmentioning

confidence: 99%

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β-Barrels and Amyloids: Structural Transitions, Biological Functions, and Pathogenesis

Sulatskaya

Kosolapova

Bobylev

et al. 2021

IJMS

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Insoluble protein aggregates with fibrillar morphology called amyloids and β-barrel proteins both share a β-sheet-rich structure. Correctly folded β-barrel proteins can not only function in monomeric (dimeric) form, but also tend to interact with one another—followed, in several cases, by formation of higher order oligomers or even aggregates. In recent years, findings proving that β-barrel proteins can adopt cross-β amyloid folds have emerged. Different β-barrel proteins were shown to form amyloid fibrils in vitro. The formation of functional amyloids in vivo by β-barrel proteins for which the amyloid state is native was also discovered. In particular, several prokaryotic and eukaryotic proteins with β-barrel domains were demonstrated to form amyloids in vivo, where they participate in interspecies interactions and nutrient storage, respectively. According to recent observations, despite the variety of primary structures of amyloid-forming proteins, most of them can adopt a conformational state with the β-barrel topology. This state can be intermediate on the pathway of fibrillogenesis (“on-pathway state”), or can be formed as a result of an alternative assembly of partially unfolded monomers (“off-pathway state”). The β-barrel oligomers formed by amyloid proteins possess toxicity, and are likely to be involved in the development of amyloidoses, thus representing promising targets for potential therapy of these incurable diseases. Considering rapidly growing discoveries of the amyloid-forming β-barrels, we may suggest that their real number and diversity of functions are significantly higher than identified to date, and represent only “the tip of the iceberg”. Here, we summarize the data on the amyloid-forming β-barrel proteins, their physicochemical properties, and their biological functions, and discuss probable means and consequences of the amyloidogenesis of these proteins, along with structural relationships between these two widespread types of β-folds.

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Section: The Transition Of β-Barrel Proteins To Amyloidsmentioning

confidence: 99%

Section: The Transition Of β-Barrel Proteins To Amyloidsmentioning

confidence: 99%

β-Barrels and Amyloids: Structural Transitions, Biological Functions, and Pathogenesis

Sulatskaya

Kosolapova

Bobylev

et al. 2021

IJMS

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“…In contrast, the D-Exon6 isoform shows a pattern of very robust protein aggregation, and, uniquely to this isoform, apparent protein ribbons. While GFP-tagged proteins have previously been reported to form aggregates [ 35 , 36 ], we posit that it is unlikely that these ribbon aggregates were solely due to overexpression or the GFP-tag, because the other isoforms did not demonstrate these ribbon formations. Other intracellular protein aggregates, such as neurofibrillary tangles and TDP43, occur in AD.…”

Section: Discussionmentioning

confidence: 90%

“…When these isoforms were transfected into HMC3 microglial cells, the expression pattern of the full-length ( Figure 6 A), D-3bp ( Figure 6 B), and D-69 ( Figure 6 C) ABI3 isoforms appeared very similar, manifesting as fine puncta with a dispersed cytosolic localization. A dispersed cytosolic localization for ABI3 fusion proteins has been described previously [ 14 ], and GFP fusion proteins often appear as similar puncta [ 35 , 36 ]. In contrast to the patterns observed with the other isoforms, the D-Exon6 ( Figure 6 D) isoform primarily manifested as unique “ribbon” formations or very large aggregates.…”

Section: Resultsmentioning

confidence: 95%

Identification and Quantitation of Novel ABI3 Isoforms Relative to Alzheimer’s Disease Genetics and Neuropathology

Turner

Shaw

Simpson

et al. 2022

Genes

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Elucidating the actions of genetic polymorphisms associated with the risk of Alzheimer’s disease (AD) may provide novel insights into underlying mechanisms. Two polymorphisms have implicated ABI3 as a modulator of AD risk. Here, we sought to identify ABI3 isoforms expressed in human AD and non-AD brain, quantify the more abundant isoforms as a function of AD genetics and neuropathology, and provide an initial in vitro characterization of the proteins produced by these novel isoforms. We report that ABI3 expression is increased with AD neuropathology but not associated with AD genetics. Single-cell RNAseq of APP/PS1 mice showed that Abi3 is primarily expressed by microglia, including disease-associated microglia. In human brain, several novel ABI3 isoforms were identified, including isoforms with partial or complete loss of exon 6. Expression of these isoforms correlated tightly with total ABI3 expression but were not influenced by AD genetics. Lastly, we performed an initial characterization of these isoforms in transfected cells and found that, while full-length ABI3 was expressed in a dispersed punctate fashion within the cytosol, isoforms lacking most or all of exon six tended to form extensive protein aggregates. In summary, ABI3 expression is restricted to microglia, is increased with Alzheimer’s neuropathology, and includes several isoforms that display a variable tendency to aggregate when expressed in vitro.

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“…As a cell trait we included copGFP and uncoupled its expression by a posttranslational self-cleavage site from our CD63-like NoMi protein which had a high EV association tendency. CopGFP was chosen over eGFP because of its quaternary structure (~104 kDa) in mammalian cells [ 64 – 67 ]. Compared to monomeric eGFP (26.9 kDa), the bulky format of copGFP reduces its chance of being packaged as a protein in NoMi-EVs, favoring its retention in the cytosol.…”

Section: Discussionmentioning

confidence: 99%

Using genetically modified extracellular vesicles as a non-invasive strategy to evaluate brain-specific cargo

Rufino-Ramos

Lüle²,

Mahjoum³

et al. 2022

Biomaterials

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New findings on GFP-like protein application as fluorescent tags: Fibrillogenesis, oligomerization, and amorphous aggregation

Cited by 16 publications

References 44 publications

β-Barrels and Amyloids: Structural Transitions, Biological Functions, and Pathogenesis

β-Barrels and Amyloids: Structural Transitions, Biological Functions, and Pathogenesis

Identification and Quantitation of Novel ABI3 Isoforms Relative to Alzheimer’s Disease Genetics and Neuropathology

Using genetically modified extracellular vesicles as a non-invasive strategy to evaluate brain-specific cargo

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