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Reliability concern: This paper was retracted on 2024-11-04. See notice: 10.1002/cbic.202400798.
Mutations in tumor suppressor genes, such as Tumor Protein 53 (TP53), are heavily implicated in aggressive cancers giving rise to gain- and loss-of-function phenotypes. While individual domains of the p53 protein have been studied extensively, structural information for full-length p53 remains incomplete. Functionalized microprocessor chips (microchips) with properties amenable to electron microscopy permitted us to visualize complete p53 assemblies for the first time. The new structures revealed p53 in an inactive dimeric state independent of DNA binding. Residues located at the protein-protein interface corresponded with modification sites in cancer-related hot spots. Changes in these regions may amplify the toxic effects of clinical mutations. Taken together, these results contribute advances in technology and imaging approaches to decode native protein models in different states of activation.
Strengths
The SiN microchip functionalization with Ni-NTA films addresses a documented limitation in cryo-EM sample preparation for small proteins (<100 kDa), demonstrated by the failure of conventional holey carbon and patterned gold grids to retain p53 particles. This provides a reproducible, commercially available alternative.
↳ Results, SiN Microchips for Cryo-EM Applications; Figure 1C-E
The inactive dimer interface residues overlap with known cancer hot spot mutations (R175, R248-249, R273, R280, R282-R283, G245, G293, E294), providing a structural rationale for mutation effects that was previously unavailable for full-length native p53.
↳ Results, Inactive P53 Assemblies Limit DNA Engagement; Figures 3B-C
The image processing workflow uses standard tools (RELION, CTFFIND-4.1) with appropriate symmetry treatment (C2 for dimer), and the molecular modeling pipeline includes MolProbity validation. The use of SDS-PAGE and western blot analysis provides orthogonal confirmation of the dimer molecular weight (~100 kDa).
↳ Experimental Section; Table 1; Figure 1B
Limitations
The claimed 4.2 Å resolution from ~8,000 particles on a 200 kV Talos F200C with a CETA (indirect) camera is at the optimistic boundary of what has been reported in the literature. Modern high-resolution cryo-EM structures typically use tens of thousands to millions of particles with direct electron detectors.
↳ Table 1; Experimental Section, EM Data Collection and Image Processing
For a 2024 structural biology paper, deposition of EM density maps and refined atomic coordinates in EMDB/PDB is a community standard expectation. The absence of any mention of deposition significantly limits independent verification of the reported structures.
↳ Experimental Section; no deposition statement present
The DNA-bound active p53 dimer model is produced by superimposing the new full-length p53 model onto the crystal structure DBD-DNA complex (PDB 2AC0), not by experimental cryo-EM determination. This limits the strength of claims about the activation mechanism and conformational transitions described in Figure 5 and Movies S5-S7.
↳ Results, Conformational Changes Support DNA Binding Events; Figure 4; Figure 5
Withdrawn — shown for reference only
This paper presents a genuine technical innovation in cryo-EM sample preparation using SiN microchips and applies it to a high-value structural target: full-length native p53 from cancer cells. The inactive dimer structure and its overlap with cancer hot spot mutations represent a novel finding, and the work is positioned within a well-established research trajectory on p53 biology. The principal tension in evaluating this work lies between the significance of the structural target and the technical limitations of the data: the small particle count (~8,000), use of an indirect CETA detector on a 200 kV instrument, and unexplained buffer discrepancy between text sections introduce meaningful uncertainty about the reported 4.2 Å resolution. The absence of EMDB/PDB data deposition, expected for a 2024 structural biology paper, further limits independent verification. The paper is best understood as a proof-of-concept that opens a productive structural direction rather than a definitive characterization of full-length p53.
Moderate3.2/5.0
The paper provides a genuine dual contribution: SiN microchip methodology for cryo-EM of small native proteins and the first full-length cryo-EM structure of native p53 in an inactive dimeric state. However, the biological novelty is constrained because the inactive dimer conformation is consistent with prior biochemical work (Fersht kinetics, co-translational dimer biogenesis) and the active state model is entirely computational.
first full-length structures of p53 proteins formed in primary brain tumors
The cryo-EM and modeling pipeline is competently described with appropriate software and validation (MolProbity), but the small particle count (~8,000) for a claimed 4.2 Å resolution on a 200 kV instrument with an indirect CETA detector is at the boundary of field standards. An unexplained buffer composition discrepancy between sections and limited detail on particle rejection and classification steps reduce methodological transparency.
The paper is well-organized with logical flow from technical challenge through methodology to structural findings and biological interpretation. Qualifying language is generally appropriate, with the activation mechanism consistently described as a 'working hypothesis,' though the abstract language is slightly stronger than the body text supports.
we have developed a working hypothesis to describe potential conformational changes
The paper substantively engages relevant p53 literature including Fersht's kinetic data, co-translational dimer biogenesis, and DBD crystal structures. However, discussion is selective and largely supportive of the authors' narrative without systematic review of existing partial structural information or deep engagement with alternative explanations for the observed dimer conformation.
Concerns4 of 4 checks
The paper is broadly self-consistent in its core structural claims, but contains a buffer composition discrepancy between sections and overclaiming in the abstract regarding 'primary brain tumors' when U87MG is an established cell line.
No data deposition in EMDB/PDB is mentioned despite 2024 community standards for structural biology, and no conflict of interest or ethics approval statements are present.
Flags: 0 declared / 5 total
All 48 cited references resolved to real publications via PubMed, Crossref, or OpenAlex.
This paper was retracted on 2024-11-04. See notice: 10.1002/cbic.202400798.
Sources: Retraction Watch ✓PubPeer (coming soon)
Low2.9/5.0
The paper addresses a well-documented, high-priority problem—full-length p53 structural characterization for cancer biology—but the connection to specific drug development programs or clinical stakeholders remains general rather than operationally specific.
significant insights for disease-related assemblies to benefit drug discovery or therapeutic opportunities
The work represents foundational structural characterization at the proof-of-concept level. The inactive dimer structure is experimentally determined but the activation mechanism and drug discovery applications require substantial downstream validation.
further biochemical investigation and future studies are aimed at delineating these steps
Evidence is limited to a single target protein (p53) from a single cancer cell line (U87MG). The SiN microchip methodology is presented as broadly applicable but requires protein-specific functionalization and has not been demonstrated across multiple targets or laboratories.
The paper connects to a rich existing evidence base spanning decades of p53 biology and fills a specific structural gap (full-length native structure). The findings are positioned to enable future work on PTM-mediated activation and mutation-specific structural effects, though several experimental steps remain before downstream applications.
Nabu’s assessment, alongside the field’s view.
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