Abstract Hereditary transthyretin amyloidosis (ATTRv) is a fatal autosomal dominant disease characterized by systemic deposition of transthyretin (TTR) amyloid fibrils, leading to progressive neuropathy and cardiomyopathy. More than 130 pathogenic mutations in the TTR gene have been identified, but their roles in TTR fibril formation and disease pathogenesis remain unclear. Here, using cryo-electron microscopy (cryo-EM), we present nineteen high-resolution TTR fibril structures (1.9-3.4 Å) from gastrocnemius muscle biopsies and vitreous humor of ten living ATTRv patients carrying nine distinct heterozygous mutations. Deep-learning-based analysis of cryo-EM densities enables semi-quantitative assessment of mutant or wild-type dominance within fibrils. These compositional profiles, combined with their structures, suggest an association between disease onset and the TTR species (wild-type or mutant) that primarily initiates amyloid formation. This biopsy-based workflow broadens access to patient tissue for amyloid structural studies, enabling systematic investigation of heterogeneous hereditary amyloidoses and the role of mutations in amyloid formation.

Leukocyte chemotactic factor 2 is a recently identified amyloidogenic protein, whose abnormal aggregation defines a systemic amyloidosis termed ALECT2 amyloidosis. Due to the lack of reliable biomarkers, diagnosis relies primarily on histological demonstration and typing of amyloid deposits in renal biopsies. However, immunohistochemical detection of ALECT2 is often inconsistent, leading to diagnostic uncertainty. The underlying basis remains poorly understood, reflecting our limited knowledge of ALECT2 deposits. Here, using cryo-electron microscopy (cryo-EM), we determined the structures of ALECT2 filaments from renal biopsies of five living patients. Unlike filaments assembled from recombinant proteins in vitro, all 133 residues of mature LECT2 are incorporated into the filament cores, with native disulfide linkages preserved. The filaments consistently adopt the shared six-layered folds in all five patients, indicating a common mechanism of amyloidogenesis. Because all residues are incorporated into the fibril core, epitope accessibility is limited. This can explain variability in immunohistochemical detection and thus highlights the need for conformation-specific antibodies and antibody-independent detection strategies for improving diagnostic accuracy. This biopsy-based workflow not only expands the availability of patient-derived tissue for cryo-EM studies but also demonstrates the potential of cryo-EM as a tool for precise diagnosis of systemic amyloidosis.

Mutations in MAPT, the tau gene, give rise to frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17), with abundant filamentous tau inclusions in brain cells. Mutations that encode missense variants of residue P301 are the most common and result in the formation of filamentous inclusions made of mutant four-repeat tau. Here we report the cryo-electron microscopy structures of tau filaments from five individuals belonging to three different families with mutation P301L and from one individual from a family with mutation P301T. A distinct three-lobed tau fold resembling the two-layered fold of Pick's disease was present in the individuals with P301L tau. Two different tau folds were found in the individual with mutation P301T, the less abundant of which was a variant of the three-lobed fold. The major P301T tau fold was V-shaped, with partial similarity to the four-layered tau folds of corticobasal degeneration and argyrophilic grain disease.

In this study, a 3-D model coupled with wall heat conduction is developed to simulate the self-pressurization process in a liquid hydrogen tank with filling ratio of 50 %. The self-pressurization curve from numerical method based on similarity principles matches well with experimental results. The results reveal an uneven and asymmetrical inner wall heat flux distribution. And the average heat leakage ratios into the vapor and liquid are estimated to be 15.72 % and 83.29 %, respectively. Further, the proportion of thermal-induced pressurization to the overall self-pressurization decreases from nearly 100 % in the initial stage to 70.32 % in the stable stage. Moreover, an overestimated pressure rise is obtained. This is because the high heat flux from the scaling process of similarity will accelerate the development of thermal stratification in the ullage and intensify local natural convection at the top cover. And this has ultimately resulted in excessive heat leakage absorption of the ullage.

In Alzheimer’s disease (AD), hyperphosphorylated tau propagates across brain regions through its seeding activity, much like prions. We assessed regional tau seeding activity and phosphorylation by RT-QuIC, western blotting (WB), and mass spectrometry in entorhinal cortex (ENT), inferior/middle/superior temporal gyri (ITG/MTG/STG), and cerebellum (CER). Tau-seeding was highest in ENT among the brain regions examined and correlated positively with Braak staging. AD-derived RT-QuIC products formed tau fibrils, propagating intracellularly in cell models. WB showed significantly higher pTau217/pTau396 in ITG vs. STG/CER. Seeding-activity correlated with WB levels of pTau217/pTau262/pTau396 (p = 0.004, p = 0.0028, p = 0.003) and Braak/Thal staging in temporal regions. Anti-tau antibodies varied in immunodepleting seeding (pTau217 > TAU5 > pTau181 > pTau396). Mass spectrometry identified 11 phospho-sites in ITG tau, while pTau231/pTau262/pTau263/pTau396 were AD-specific. Regional tau-seeding correlates with local phospho-tau levels and pathology, supporting plasma pTau217 for diagnosis and seeding activity as a potential biomarker.

Access to structurally-defined human proteoforms is essential to the biochemical studies on human health and medicine. Chemical protein synthesis provides a bottom-up and atomic-resolution approach for the preparation of homogeneous proteoforms bearing any number of post-translational modifications of any structure, at any position, and in any combination. In this review, we summarize the development of chemical protein synthesis, focusing on the recent advances in synthetic methods, product characterizations, and biomedical applications. By analyzing the chemical protein synthesis studies on human proteoforms reported to date, this review demonstrates the significant methodological improvements that have taken place in the field of human proteoform synthesis, especially in the last decade. Our analysis shows that although further method development is needed, all the human proteoforms could be within reach in a cost-effective manner through a divide-and-conquer chemical protein synthesis strategy. The synthetic proteoforms have been increasingly used to support biomedical research, including spatial-temporal studies and interaction network analysis, activity quantification and mechanism elucidation, and the development and evaluation of diagnostics and therapeutics.

Mutations in MAPT, the microtubule-associated protein tau gene, give rise to cases of frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) with abundant filamentous tau inclusions in brain cells. Individuals with pathological MAPT variants exhibit behavioural changes, cognitive impairment and signs of parkinsonism. Missense mutations of residue P301, which are the most common MAPT mutations associated with FTDP-17, give rise to the assembly of mutant four-repeat tau into filamentous inclusions, in the absence of extracellular deposits. Here we report the cryo-EM structures of tau filaments from five individuals belonging to three unrelated families with mutation P301L and from one individual belonging to a family with mutation P301T. A novel three-lobed tau fold resembling the two-layered tau fold of Pick's disease was present in all cases with the P301L tau mutation. Two different tau folds were found in the case with mutation P301T, the less abundant of which was a variant of the three-lobed fold. The major P301T tau fold was V-shaped, with partial similarity to the four-layered tau folds of corticobasal degeneration and argyrophilic grain disease. These findings suggest that FTDP-17 with mutations in P301 should be considered distinct inherited tauopathies and that model systems with these mutations should be used with caution in the study of sporadic tauopathies.

A 21-nucleotide duplication in one allele of SNCA was identified in a previously described disease with abundant α-synuclein inclusions that we now call juvenile-onset synucleinopathy (JOS). This mutation translates into the insertion of MAAAEKT after residue 22 of α-synuclein, resulting in a protein of 147 amino acids. Both wild-type and mutant proteins were present in sarkosyl-insoluble material that was extracted from frontal cortex of the individual with JOS and examined by electron cryo-microscopy. The structures of JOS filaments, comprising either a single protofilament, or a pair of protofilaments, revealed a new α-synuclein fold that differs from the folds of Lewy body diseases and multiple system atrophy (MSA). The JOS fold consists of a compact core, the sequence of which (residues 36–100 of wild-type α-synuclein) is unaffected by the mutation, and two disconnected density islands (A and B) of mixed sequences. There is a non-proteinaceous cofactor bound between the core and island A. The JOS fold resembles the common substructure of MSA Type I and Type II dimeric filaments, with its core segment approximating the C-terminal body of MSA protofilaments B and its islands mimicking the N-terminal arm of MSA protofilaments A. The partial similarity of JOS and MSA folds extends to the locations of their cofactor-binding sites. In vitro assembly of recombinant wild-type α-synuclein, its insertion mutant and their mixture yielded structures that were distinct from those of JOS filaments. Our findings provide insight into a possible mechanism of JOS fibrillation in which mutant α-synuclein of 147 amino acids forms a nucleus with the JOS fold, around which wild-type and mutant proteins assemble during elongation.

Carotenoid (Car) pigments perform central roles in photosynthesis-related light harvesting (LH), photoprotection, and assembly of functional pigment-protein complexes. However, the relationships between Car depletion in the LH, assembly of the prokaryotic reaction center (RC)-LH complex, and quinone exchange are not fully understood. Here, we analyzed native RC-LH (nRC-LH) and Car-depleted RC-LH (dRC-LH) complexes in Roseiflexus castenholzii, a chlorosome-less filamentous anoxygenic phototroph that forms the deepest branch of photosynthetic bacteria. Newly identified exterior Cars functioned with the bacteriochlorophyll B800 to block the proposed quinone channel between LHαβ subunits in the nRC-LH, forming a sealed LH ring that was disrupted by transmembrane helices from cytochrome c and subunit X to allow quinone shuttling. dRC-LH lacked subunit X, leading to an exposed LH ring with a larger opening, which together accelerated the quinone exchange rate. We also assigned amino acid sequences of subunit X and two hypothetical proteins Y and Z that functioned in forming the quinone channel and stabilizing the RC-LH interactions. This study reveals the structural basis by which Cars assembly regulates the architecture and quinone exchange of bacterial RC-LH complexes. These findings mark an important step forward in understanding the evolution and diversity of prokaryotic photosynthetic apparatus.

The hepatitis B virus core protein (HBcAg) is a highly immunogenic particulate antigen. Nearly all patients with persistent or resolved hepatitis B virus (HBV) infection show seropositivity for hepatitis B core antibody (anti-HBc), which appears in the early stage of infection and is mostly present for life. Traditionally, the anti-HBc is regarded as an evidential serological marker of HBV infections. In the last ten years, several studies revealed the predictive value of quantitative anti-HBc (qAnti-HBc) level in the treatment response and clinical outcome of chronic HBV infections, implying new insights into this classic marker. Overall, qAnti-HBc should be regarded as an indicator of the host’s immune response specific to HBV, which correlates with HBV-related hepatitis activity and liver pathology. This review summarized the latest understanding of the clinical values of qAnti-HBc for differentiating the CHB phase, predicting treatment response, and providing disease prognosis. Moreover, we also discussed the possible mechanism of qAnti-HBc regulation during different courses of HBV infection.

Positron emission tomography (PET) imaging allows monitoring the progression of amyloid aggregation in the living brain. [18F]-Flortaucipir is the only approved PET tracer compound for the visualisation of tau aggregation. Here, we describe cryo-EM experiments on tau filaments in the presence and absence of flortaucipir. We used tau filaments isolated from the brain of an individual with Alzheimer's disease (AD), and from the brain of an individual with primary age-related tauopathy (PART) with a co-pathology of chronic traumatic encephalopathy (CTE). Unexpectedly, we were unable to visualise additional cryo-EM density for flortaucipir for AD paired helical or straight filaments (PHFs or SFs), but we did observe density for flortaucipir binding to CTE Type I filaments from the case with PART. In the latter, flortaucipir binds in a 1:1 molecular stoichiometry with tau, adjacent to lysine 353 and aspartate 358. By adopting a tilted geometry with respect to the helical axis, the 4.7 Å distance between neighbouring tau monomers is reconciled with the 3.5 Å distance consistent with π-π-stacking between neighbouring molecules of flortaucipir.

The amyotrophic lateral sclerosis/parkinsonism-dementia complex (ALS/PDC) of the island of Guam and the Kii peninsula of Japan is a fatal neurodegenerative disease of unknown cause that is characterized by the presence of abundant filamentous tau inclusions in brains and spinal cords. Here, we used electron cryo-microscopy to determine the structures of tau filaments from the cerebral cortex of three cases of ALS/PDC from Guam and eight cases from Kii, as well as from the spinal cord of two of the Guam cases. Tau filaments had the chronic traumatic encephalopathy (CTE) fold, with variable amounts of Type I and Type II filaments. Paired helical tau filaments were also found in three Kii cases and tau filaments with the corticobasal degeneration fold in one Kii case. We identified a new Type III CTE tau filament, where protofilaments pack against each other in an antiparallel fashion. ALS/PDC is the third known tauopathy with CTE-type filaments and abundant tau inclusions in cortical layers II/III, the others being CTE and subacute sclerosing panencephalitis. Because these tauopathies are believed to have environmental causes, our findings support the hypothesis that ALS/PDC is caused by exogenous factors.

Many age-dependent neurodegenerative diseases, such as Alzheimer’s and Parkinson’s, are characterized by abundant inclusions of amyloid filaments. Filamentous inclusions of the proteins tau, amyloid-β, α-synuclein and transactive response DNA-binding protein (TARDBP; also known as TDP-43) are the most common 1 , 2 . Here we used structure determination by cryogenic electron microscopy to show that residues 120–254 of the lysosomal type II transmembrane protein 106B (TMEM106B) also form amyloid filaments in human brains. We determined the structures of TMEM106B filaments from a number of brain regions of 22 individuals with abundant amyloid deposits, including those resulting from sporadic and inherited tauopathies, amyloid-β amyloidoses, synucleinopathies and TDP-43 proteinopathies, as well as from the frontal cortex of 3 individuals with normal neurology and no or only a few amyloid deposits. We observed three TMEM106B folds, with no clear relationships between folds and diseases. TMEM106B filaments correlated with the presence of a 29-kDa sarkosyl-insoluble fragment and globular cytoplasmic inclusions, as detected by an antibody specific to the carboxy-terminal region of TMEM106B. The identification of TMEM106B filaments in the brains of older, but not younger, individuals with normal neurology indicates that they form in an age-dependent manner.

Parkinson’s disease (PD) is the most common movement disorder, with resting tremor, rigidity, bradykinesia and postural instability being major symptoms (1). Neuropathologically, it is characterised by the presence of abundant filamentous inclusions of α-synuclein in the form of Lewy bodies and Lewy neurites in some brain cells, including dopaminergic nerve cells of the substantia nigra (2). PD is increasingly recognised as a multisystem disorder, with cognitive decline being one of its most common non-motor symptoms. Many patients with PD develop dementia more than 10 years after diagnosis (3). PD dementia (PDD) is clinically and neuropathologically similar to dementia with Lewy bodies (DLB), which is diagnosed when cognitive impairment precedes parkinsonian motor signs or begins within one year from their onset (4). In PDD, cognitive impairment develops in the setting of well-established PD. Besides PD and DLB, multiple system atrophy (MSA) is the third major synucleinopathy (5). It is characterised by the presence of abundant filamentous α-synuclein inclusions in brain cells, especially oligodendrocytes (Papp-Lantos bodies). We previously reported the electron cryo-microscopy (cryo-EM) structures of two types of α-synuclein filaments extracted from the brains of individuals with MSA (6). Each filament type is made of two different protofilaments. Here we report that the cryo-EM structures of α-synuclein filaments from the brains of individuals with PD, PDD and DLB are made of a single protofilament (Lewy fold) that is markedly different from the protofilaments of MSA. These findings establish the existence of distinct molecular conformers of assembled α-synuclein in neurodegenerative disease.

Mutations in the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike receptor-binding domain (RBD) may alter viral host tropism and affect the activities of neutralizing antibodies. Here, we investigated 153 RBD mutants and 11 globally circulating variants of concern (VOCs) and variants of interest (VOIs) (including Omicron) for their antigenic changes and cross-species tropism in cells expressing 18 ACE2 orthologs. Several RBD mutations strengthened viral infectivity in cells expressing ACE2 orthologs of non-human animals, particularly those less susceptible to the ancestral strain. The mutations surrounding amino acids (aas) 439-448 and aa 484 are more likely to cause neutralization resistance. Strikingly, enhanced cross-species infection potential in the mouse and ferret, instead of the neutralization-escape scores of the mutations, account for the positive correlation with the cumulative prevalence of mutations in humans. These findings present insights for potential drivers of circulating SARS-CoV-2 variants and provide informative parameters for tracking and forecasting spreading mutations.

In recent years, dimethylphenol has played an important role in industrial production. The demand and application of dimethylphenol will increase greatly with the continuous advancement of the internationalization process. Solid-liquid equilibrium (SLE) is fundamental for the separation process design, the SLE data of dimethylphenol plays an important role in designing the melting and crystallization separation process. The SLE data of mixtures of 2,6-dimethylphenol (2,6-DMP), 3,5-dimethylphenol (3,5-DMP) and 2,5-dimethylphenol (2,5-DMP) were determined by differential scanning calorimetry (DSC). The crystallographic analysis of eutectic mixtures was analyzed by FTIR. The experimental results show that all the systems studied in this paper exhibit eutectic behavior. SLE data was correlated with Wilson and NRTL models and a satisfactory correlation was obtained. The study provided reliable and accurate data for separating the mixtures of 2,6-dimethylphenol, 3,5-dimethylphenol and 2,5-dimethylphenol. These SLE data are of great significance for the process development and process optimization of dimethylphenol.

EMPIAR, the Electron Microscopy Public Image Archive centered at EMBL-EBI, is a public resource for raw electron microscopy images related to EMDB, contains micrographs, particle sets and tilt-series.

Parkinson's disease (PD) is the most common movement disorder, with resting tremor, rigidity, bradykinesia and postural instability being major symptoms1. Neuropathologically, it is characterized by the presence of abundant filamentous inclusions of α-synuclein in the form of Lewy bodies and Lewy neurites in some brain cells, including dopaminergic nerve cells of the substantia nigra2. PD is increasingly recognised as a multisystem disorder, with cognitive decline being one of its most common non-motor symptoms. Many patients with PD develop dementia more than 10 years after diagnosis3. PD dementia (PDD) is clinically and neuropathologically similar to dementia with Lewy bodies (DLB), which is diagnosed when cognitive impairment precedes parkinsonian motor signs or begins within one year from their onset4. In PDD, cognitive impairment develops in the setting of well-established PD. Besides PD and DLB, multiple system atrophy (MSA) is the third major synucleinopathy5. It is characterized by the presence of abundant filamentous α-synuclein inclusions in brain cells, especially oligodendrocytes (Papp-Lantos bodies). We previously reported the electron cryo-microscopy structures of two types of α-synuclein filament extracted from the brains of individuals with MSA6. Each filament type is made of two different protofilaments. Here we report that the cryo-electron microscopy structures of α-synuclein filaments from the brains of individuals with PD, PDD and DLB are made of a single protofilament (Lewy fold) that is markedly different from the protofilaments of MSA. These findings establish the existence of distinct molecular conformers of assembled α-synuclein in neurodegenerative disease.