Molecular Mechanisms Of Assembly Research Articles

Translocation of subunit PPSE in plipastatin synthase and synthesis of novel lipopeptides

Nonribosomal peptide synthase (NRPS) is a unique molecular assembly mechanism with high hybridity. Its recombination is conducive to the development of novel lipopeptides. However, there are few reports on NRPS subunit recombination of plipastatin at present. In this paper, plipastatin synthase was modified by the forward movement of subunit PPSE and the replacement of the communication-mediating (COM) domain. The results showed that ppsABE, a new assembly line, could synthesize novel lipopeptides such as cycle pentapeptide (C16-18β-OHFA-E-O-cyclo(Y-T-I), and its antimicrobial activity against Rhizopus stolonifer and Staphylococcus aureus was better than that of plipastatin. However, the reactivity of ppsABCE disappeared, but the substitution of COMDppsC/COMAppsD or COMDppsD/COMAppsE for COMDppsC/COMAppsE could restore its activity and conduct the biosynthesis of linear hexapeptide (C16-17β-OHFA-E-O-Y-T-E-A/V) and heptapeptide (C17-18β-OHFA-E-O-Y-T-E-A-I). Collectively, these findings indicated that the COM donor domain at the C-terminus of PPSB could communicate with the COM acceptor domain at the N-terminus of PPSE and that the compatible COM domain is an important tool for communication between nonpartner subunits. Moreover, the integrity and selective compatibility of the COM acceptor domain of subunit PPSE are essential to promote the interaction between PPSE and other subunits. This work further complemented the rules of NRPS subunit recombination and provided a theoretical basis for the development of novel high-efficiency lipopeptides.

The p97 segregase cofactor Ubxn7 facilitates replisome disassembly during S-phase

Complex cellular processes are driven by the regulated assembly and disassembly of large multiprotein complexes. While we are beginning to understand the molecular mechanism for assembly of the eukaryotic DNA replication machinery (replisome), we still know relatively little about the regulation of its disassembly at replication termination. Recently, the first elements of this process have emerged, revealing that the replicative helicase, at the heart of the replisome, is polyubiquitylated prior to unloading and that this unloading requires p97 segregase activity. Two different E3 ubiquitin ligases have now been shown to ubiquitylate the helicase under different conditions: Cul2Lrr1 and TRAIP. Here, using Xenopus laevis egg extract cell-free system and biochemical approaches, we have found two p97 cofactors, Ubxn7 and Faf1, which can interact with p97 during replisome disassembly during S-phase. We show only Ubxn7, however, facilitates efficient replisome disassembly. Ubxn7 delivers this role through its interaction via independent domains with both Cul2Lrr1 and p97 to allow coupling between Mcm7 ubiquitylation and its removal from chromatin. Our data therefore characterize Ubxn7 as the first substrate-specific p97 cofactor regulating replisome disassembly in vertebrates and a rationale for the efficacy of the Cul2Lrr1 replisome unloading pathway in unperturbed S-phase.

Mre11-Rad50 oligomerization promotes DNA double-strand break repair

The conserved Mre11-Rad50 complex is crucial for the detection, signaling, end tethering and processing of DNA double-strand breaks. While it is known that Mre11-Rad50 foci formation at DNA lesions accompanies repair, the underlying molecular assembly mechanisms and functional implications remained unclear. Combining pathway reconstitution in electron microscopy, biochemical assays and genetic studies, we show that S. cerevisiae Mre11-Rad50 with or without Xrs2 forms higher-order assemblies in solution and on DNA. Rad50 mediates such oligomerization, and mutations in a conserved Rad50 beta-sheet enhance or disrupt oligomerization. We demonstrate that Mre11-Rad50-Xrs2 oligomerization facilitates foci formation, DNA damage signaling, repair, and telomere maintenance in vivo. Mre11-Rad50 oligomerization does not affect its exonuclease activity but drives endonucleolytic cleavage at multiple sites on the 5′-DNA strand near double-strand breaks. Interestingly, mutations in the human RAD50 beta-sheet are linked to hereditary cancer predisposition and our findings might provide insights into their potential role in chemoresistance.

Co-operative binding of SKP1, Cullin1 and Cullin7 to FBXW8 results in Cullin1-SKP1-FBXW8-Cullin7 functional complex formation that monitors cellular function of β-TrCP1

F-box protein FBXW8 is known to interact with scaffolding protein Cullin1 and Cullin7 to form SCF (SKP1, Cullin and F-box protein) complex. However, detail understanding about the importance of both Cullins for SCF-FBXW8 complex formation as well as its ubiquitin ligase activity remains elusive. Here, we show that, through in vitro and in vivo studies, Cullin1 and Cullin7 increase each other's binding to FBXW8 synergistically. Interestingly, absence of either Cullin results in abrogation of binding of other Cullin to FBXW8. Binding of SKP1 to FBXW8 also increases in the presence of both the Cullins. Thus, SKP1, Cullin1 and Cullin7 are essential to form Cullin1-SKP1-FBXW8-Cullin7 functional ubiquitin ligase complex. Further, using computational, mutational and biochemical analysis, we found that Cullin1 binds to N-terminus of FBXW8 through SKP1 while Cullin7 associates with C-terminus of FBXW8 to form Cullin1-SKP1-FBXW8-Cullin7 functional complex in a cooperative manner. Results showed that Cullin1-SKP1-FBXW8-Cullin7 complex plays a key role in maintaining the basal level expression of β-TrCP1. Moreover, Cullin1-SKP1-FBXW8-Cullin7 complex promotes cell migration by activating β-catenin via directing proteasomal degradation of β-TrCP1. Overall, our study reveals the intriguing molecular mechanism of assembly of SKP1, Cullin1, Cullin7 and FBXW8 to form Cullin1-SKP1-FBXW8-Cullin7 functional complex that control the function of β-TrCP1.

Molecular mechanism of calcium induced trimerization of C1q-like domain of otolin-1 from human and zebrafish

The C1q superfamily includes proteins involved in innate immunity, insulin sensitivity, biomineralization and more. Among these proteins is otolin-1, which is a collagen-like protein that forms a scaffold for the biomineralization of inner ear stones in vertebrates. The globular C1q-like domain (gC1q), which is the most conserved part of otolin-1, binds Ca2+ and stabilizes its collagen-like triple helix. The molecular details of the assembly of gC1q otolin-1 trimers are not known. Here, we substituted putative Ca2+-binding acidic residues of gC1q otolin-1 with alanine to analyse how alanine influences the formation of gC1q trimers. We used human and zebrafish gC1q otolin-1 to assess how evolutionary changes affected the function of the protein. Surprisingly, the mutated forms of gC1q otolin-1 trimerized even in the absence of Ca2+, although they were less stable than native proteins saturated with Ca2+. We also found that the zebrafish gC1q domain was less stable than the human homologue under all tested conditions and became stabilized at higher concentrations of Ca2+, which showed that specific interactions leading to the neutralization of the negative charge at the axis of a gC1q trimer by Ca2+ are required for the trimers to form. Moreover, human gC1q otolin-1 seems to be optimized to function at lower concentrations of Ca2+, which is consistent with reported Ca2+ concentrations in the endolymphs of fish and mammals. Our results allow us to explain the molecular mechanism of assembly of proteins from the C1q superfamily, the modulating role of Ca2+ and expand the knowledge of biomineralization of vertebrate inner ear stones: otoliths and otoconia.

Putative hexameric glycosyltransferase functional unit revealed by the crystal structure of Acinetobacter baumannii MurG.

Lipid II, the main component of the bacterial cell wall, is synthesized by the addition of UDP-N-acetylglucosamine to the UDP-N-acetylmuramic acid pentapeptide catalyzed by the glycosyltransferase MurG. Owing to its critical role in cell-wall biosynthesis, MurG is considered to be an attractive target for antibacterial agents. Although the Mur family ligases have been extensively studied, the molecular mechanism of the oligomeric scaffolding assembly of MurG remains unclear. In this study, MurG from Acinetobacter baumannii (abMurG), a human pathogen, was characterized and its hexameric crystal structure was unveiled; this is the first homo-oligomeric structure to be described in the MurG family and the Mur family. Homogeneous protein samples were produced for structural studies using size-exclusion chromatography, the absolute molecular mass was calculated via multi-angle light scattering, and protein-protein interactions were analyzed using the PDBePISA server. abMurG was found to form homo-oligomeric complexes in solution, which might serve as functional units for the scaffolding activity of MurG. Furthermore, analysis of this structure revealed the molecular assembly mechanism of MurG. This structural and biochemical study elucidated the homo-oligomerization mechanism of MurG and suggests a new potential antibiotic target on MurG.

Molecular mechanisms of assembly and TRIP13-mediated remodeling of the human Shieldin complex

The Shieldin complex, composed of REV7, SHLD1, SHLD2, and SHLD3, protects DNA double-strand breaks (DSBs) to promote nonhomologous end joining. The AAA+ ATPase TRIP13 remodels Shieldin to regulate DNA repair pathway choice. Here we report crystal structures of human SHLD3-REV7 binary and fused SHLD2-SHLD3-REV7 ternary complexes, revealing that assembly of Shieldin requires fused SHLD2-SHLD3 induced conformational heterodimerization of open (O-REV7) and closed (C-REV7) forms of REV7. We also report the cryogenic electron microscopy (cryo-EM) structures of the ATPγS-bound fused SHLD2-SHLD3-REV7-TRIP13 complexes, uncovering the principles underlying the TRIP13-mediated disassembly mechanism of the Shieldin complex. We demonstrate that the N terminus of REV7 inserts into the central channel of TRIP13, setting the stage for pulling the unfolded N-terminal peptide of C-REV7 through the central TRIP13 hexameric channel. The primary interface involves contacts between the safety-belt segment of C-REV7 and a conserved and negatively charged loop of TRIP13. This process is mediated by ATP hydrolysis-triggered rotatory motions of the TRIP13 ATPase, thereby resulting in the disassembly of the Shieldin complex.

Roles of type IIa receptor protein tyrosine phosphatases as synaptic organizers.

Neurons establish circuits for brain functions such as cognition, emotion, learning, and memory. Their connections are mediated by synapses, which are specialized cell-cell adhesions responsible for neuronal signal transmission. During neurodevelopment, synapse formation is triggered by interactions of cell adhesion molecules termed synaptic organizers or synapse organizers. Type IIa receptor protein tyrosine phosphatases (IIa RPTPs; also known as leukocyte common antigen-related receptor tyrosine phosphatases or LAR-RPTPs) play important roles in axon guidance and neurite extension, and also serve as presynaptic organizers. IIa RPTPs transsynaptically interact with multiple sets of postsynaptic organizers, mostly in a splicing-dependent fashion. Here, we review and update research progress on IIa RPTPs, particularly regarding their functional roles in vivo demonstrated using conditional knockout approach and structural insights into their extracellular and intracellular molecular interactions revealed by crystallography and other biophysical techniques. Future directions in the research field of IIa RPTPs are also discussed, including recent findings of the molecular assembly mechanism underlying the formation of synapse-specific nanostructures essential for synaptic functions.

Amyloid-like peptide nanofibrils as scaffolds for tissue engineering: Progress and challenges (Review)

Networks of amyloid-like nanofibrils assembled from short peptide sequences have the ability to form scaffolds that can encapsulate clinically relevant stem cells encouraging their attachment, growth, and differentiation into various lineages which can be used in tissue engineering applications to treat a range of diseases and traumas. In this review, the author highlights a selection of important proof-of-principle papers that show how this class of self-assembled networks is highly suited to biomaterial scaffold development. The author highlights recent studies which have shown that these scaffolds can be used to promote cell and tissue regeneration both in vitro and in vivo. The author also presents some fundamental knowledge gaps which are preventing the widespread translation of such scaffolds. Finally, the author outlines a selection of studies that elucidate molecular assembly mechanisms and biophysical properties of amyloid-like peptide nanofibrils and suggests how studies like these might lead to the ability to generate nanofibril scaffolds with bespoke properties for tissue engineering.

Co-Transcriptional Ribosome Assembly in Real-Time

The bacterial ribosome is a large macromolecular machine composed of three large RNA molecules and more than 50 proteins, which are co-transcriptionally assembled in a living cell within 1-2 minutes.The molecular mechanism of assembly has extensively been studied with pre-transcribed ribosomal RNA (rRNA) mainly using ensemble methods, in which binding of each ribosomal protein (r-protein) is detected as the average state of all populations in which it is present. Because active transcription at the correct speed is essential for ribosome assembly at physiological conditions, the molecular mechanism of ribosome assembly has to be understood co-transcriptionally.Here we present a novel single-molecule fluorescence based approach in which transcription and folding of single rRNA molecules and the binding of r-proteins to them can be visualized simultaneously in real-time.Using zero-mode waveguide instrumentation, we could monitor transcription progression of single RNA molecules of several hundred nucleotides.Studying the co-transcriptional binding of the primary r-protein S15 to the central domain shows that besides short lived binding events (few seconds lifetime), specific S15 binding (>30 seconds lifetime) can occur shortly after transcription of its binding site. In contrast, co-transcriptional binding of primary protein S7 to the entire 3’-domain is much more dynamic with some RNA molecules being unable to stably bind S7 (seconds lifetime) while others can eventually bind S7 specifically, yet can do so only minutes after transcription at 20 degrees Celsius.Overall, using a novel approach to observe in real-time binding of single r-proteins to transcribing r-RNA, we see that co-transcriptional folding of the central 16S rRNA domain is efficient allowing immediate protein binding, while 3’-domain folding is inefficient, indicating that other r-proteins or assembly factors such as RNA helicases must increase folding efficiency in the cell (at least at low temperature).

Facile fabrication of Sudan red particle microcapsules by a polymerizable gemini surfactant and molecular assembly mechanisms.

An efficient method was developed to encapsulate water insoluble organic particles of Sudan red III (SR) in aqueous suspensions by using a polymerizable cationic gemini surfactant, 1,3-bis(N,N-dimethyl-N-cetylammonium)-2-(propylacrylate dibromide) (AGC16). The AGC16 coated SR microcapsules (AGC16@SR) were prepared by absorption of AGC16 on the surface of SR, followed by in situ homopolymerization (PAGC16). Several measurements, including transmission and scanning electron microscopy, isothermal titration calorimetry, zeta potential, electron paramagnetic resonance and small angle X-ray scattering, were performed to determine the adsorption amount of AGC16, and the layer structures and the molecular assembly mechanism in the AGC16@SR and PAGC16@SR systems, respectively. For comparison purposes, the polymerizable cationic surfactant with one head group and a single alkyl chain, acryloyloxyethyl-N,N-dimethyl-N-cetylammonium bromide (referred to as ASC16), as well as the systems of ASC16@SR and PASC16@SR were also investigated in parallel. It was found that AGC16 molecules and their aggregates were simultaneously assembled into a shell layer, in which the saturated adsorption amount of AGC16 on SR is less than 1/2 that of ASC16, but the assembly layer of AGC16 is more hydrophobic with greater packing tightness compared with that of ASC16. It was also revealed that after in situ homopolymerization, the microcapsule shell becomes more compact. In the case of PAGC16@SR, the layers show higher surface roughness and irregularity compared with that of PASC16@SR. Moreover, the sustained release behavior of SR was also evaluated. The results revealed that PAGC16@SR performed well for SR controlled release, which was sorted by release performance as the following sequence: PAGC16@SR > AGC16@SR > PASC16@SR > ASC16@SR. Thus, the polymerizable cationic gemini surfactant holds substantial potential to be developed as an ideal candidate of soft matter to construct efficient controllable release systems.

Editorial: Role of lipids in virus assembly.

Editorial: Role of lipids in virus assembly.

Solvent Effects on Kinetic Mechanisms of Self-Assembly by Peptide Amphiphiles via Molecular Dynamics Simulations

Peptide amphiphiles are known to form a variety of distinctive self-assembled nanostructures (including cylindrical nanofibers in hydrogels) dependent upon the solvent conditions. Using a novel coarse-grained model, large-scale molecular dynamics simulations are performed on a system of 800 peptide amphiphiles (sequence, palmitoyl-Val3Ala3Glu3) to elucidate kinetic mechanisms of molecular assembly as a function of the solvent conditions. The assembly process is found to occur via a multistep process with transient intermediates that ultimately leads to the stabilized nanostructures including open networks of β-sheets, cylindrical nanofibers, and elongated micelles. Different kinetic mechanisms are compared in terms of peptide secondary structures, solvent-accessible surface area, radius of gyration, relative shape anisotropy, intra/intermolecular interactions, and aggregate size dynamics to provide insightful information for the design of functional biomaterials.

Bottom-up graphene-nanoribbon fabrication reveals chiral edges and enantioselectivity.

We produce precise chiral-edge graphene nanoribbons on Cu{111} using self-assembly and surface-directed chemical reactions. We show that, using specific properties of the substrate, we can change the edge conformation of the nanoribbons, segregate their adsorption chiralities, and restrict their growth directions at low surface coverage. By elucidating the molecular-assembly mechanism, we demonstrate that our method constitutes an alternative bottom-up strategy toward synthesizing defect-free zigzag-edge graphene nanoribbons.

Combining X-ray scattering and spectroscopy as structure resolving tool

Modern applications and basic research in medicine, biotechnology and materials are often concerned with hybrid (organic-inorganic) and synergetic systems. In other words, systems that brings enhanced properties and performances. The challenge is now in the understanding of the complex interactions leading to their assembly and operation. Due to their inherent chemical and structural complexities a combination of several techniques is necessary to determine unambiguously the molecular mechanisms of assembly and operation. [1, 2] To address this need, we have implemented a simultaneous measurement platform called SURF [3] that consists of SAXS, UV-Vis, Raman and fluorescence techniques. The SURF platform provides simultaneous measurements on the same sample volume and a multivariate framework to associate the spectroscopic and X-ray scattering information. Convex constraint analysis (CCA) and two dimensional correlation analyses (2DCOS and 2DHCOS) had been introduced to enhance the interpretation and integration of the data from the different techniques producing self-consistent models and resolving complex behaviour details (structure and chemistry). Additional benefits of the SURF are sample quality control and "on line" diagnostics. In this contribution, we illustrate the benefits of the SURF approach on selected examples. Acknowledgements: The SURF set-up has been mainly financial supported by MAX IV laboratory. S. Haas has a postdoctoral grant of MAX IV lab. S Canton and Q. Zhu are acknowledged for fruitful discussions.

Novel Supramolecular Thixotropic Metallohydrogels Consisting of Rare Metal–Organic Nanoparticles: Synthesis, Characterization, and Mechanism of Aggregation

Even without obvious sticky sites, novel supramolecular thixotropic metallohydrogels consisting of rare metal–organic nanoparticles (MNPs) have been readily accessible from simple structured pincer-type terpyridine Cu(II) complexes at the gelator concentration as low as 0.25 wt %. The obtained soft materials have been fully characterized by using a combination of experimental techniques including scanning electron microscopy (SEM), transmission electron microscopy (TEM), atomic force microscopy (AFM), dynamic light scattering (DLS), X-ray diffraction (XRD), high-resolution mass spectrometry (HR-MS), and rheology measurements. Based on these studies, hypothesized molecular assembly mechanisms for distinctly different morphologies observed in different solvents were proposed. Besides general heating–cooling gel preparation procedures, surprisingly, the metallohydrogel was formed by simply stirring the mixture of pincer ligand and Cu salts in water directly, further indicating thixotropic property and self-h...

Biosynthetic assembly of cytochalasin backbone

Cytochalasins are an important class of fungal natural products in view of structural diversity and biological activities. Although their biosynthetic studies have been examined extensively, the detailed molecular assembly mechanism remains to be solved. We have succeeded to heterologously express the cytochalasin polyketide synthase–non-ribosomal peptide synthetase (PKS–NRPS) hybrid gene ccsA and the trans-acting enoyl-CoA reductase gene ccsC in Aspergillus oryzae. The resultant transformant produced a novel metabolite possessing the cytochalasin backbone. This established that CcsA is capable of constructing the octaketide connected with phenylalanine in collaboration with CcsC, and that CcsA R domain catalyzes reductive cleavage of the thio-tethered PKS–NRPS product.

Heat-induced fibril assembly of vicilin at pH 2.0: Reaction kinetics, influence of ionic strength and protein concentration, and molecular mechanism

Although the heat-induced fibril assembly of legume 7S/11S globulins at pH2.0 has been reported, the kinetics and molecular mechanism of the assembly are still not fully understood. In this paper, the reaction kinetics, and dependence of fibril assembly on protein concentration and ionic strength, as well as the changes in fibril morphology of three selected vicilins (from kidney, mung and red beans; denoted as Vk, Vm and Vr, respectively) were investigated and compared by dynamic laser scattering, Thioflavin T fluorescence, and atomic force microscopy. The polypeptide hydrolysis, zeta-potential, and conformation changes during the assembly were also characterized, with the aim to unravel the molecular mechanism of the assembly. The results indicated that Vk exhibited a much higher ability to form highly ordered fibrils than Vm (or Vr), though among the three vicilins, Vm exhibited the highest potential. The differences were closely associated with the differences in their susceptibility of the polypeptides to acid hydrolysis, and more basically, amino acid composition. The amount of the formed protofibrils or fibrils in all the vicilins progressively increased with increasing protein concentration, but their size seemed to be unaffected. The fibril assembly of Vk could be greatly improved by electrostatic screening, in an ionic strength dependent way, while that of the other two vicilins was only slightly affected. This further confirmed that Vk exhibited a much better ability to assemble into fibrils. The fibril assembly of vicilins involves at least four steps: thermal denaturation and aggregation (to form amorphous aggregates), polypeptide hydrolysis of the aggregates, initiation of fibril assembly (or formation of protofibrils), and fibril elongation (to form highly ordered fibrils) or chain termination. The findings would be of great importance for understanding the molecular mechanism of fibril assembly of oligomeric proteins from plant sources.

Amyloidogenic Protein of α-Synuclein

Amyloidogenesis is the key pathological phenomenon commonly observed in various neurodegenerative disorders. α-Synuclein is the major constituent of Lewy bodies as a common pathological signature of Lewy body diseases (LBDs) including Parkinson’s disease (PD), PD with dementia (PDD), and Dementia with Lewy bodies (DLB). As proteins unfold, they would result in producing either ordered or disordered aggregates unless they are folded back to the native state by molecular chaperones or removed via proteolytic degradation. α-Synuclein known as a natively unfolded protein has self-assembled into the ordered protein aggregates of amyloid fibrils which comprise the radiating filaments found in Lewy bodies. Amyloid fibrils are generated via either a template-dependent or template-independent mechanism. The prevalent nucleation-dependent fibrillation accelerates the assembly process in the presence of seeds such as prefibrillar species. As a template-independent process, we have recently proposed the double-concerted fibrillation mechanism in which the oligomeric species of α-synuclein act as a growing unit to form the mature fibrils. Despite insufficient understanding of the toxic causes of α-synuclein, the oligomeric species have been suggested to be responsible for the cellular degeneration by influencing membrane stability while leaving the amyloid fibrils as a detoxification end product. Alternatively, the transition process from the oligomers to the fibrils has been proposed to affect cell viability. It is, therefore, expected to develop prophylactic and therapeutic strategies toward the synucleinopathies by studying cellular function of α-synuclein, molecular mechanism of its assembly into the amyloid fibrils, and their effects on cellular biogenesis. By studying cellular function of α-synuclein, its molecular mechanism of assembly into amyloid fibrils and their effects on cellular biogenesis, progress of prophylactic and therapeutic strategies toward synucleinopathy can be seen.

On the Mechanism of αC Polymer Formation in Fibrin

Our previous studies revealed that the fibrinogen αC-domains undergo conformational changes and adopt a physiologically active conformation upon their self-association into αC polymers in fibrin. In the present study, we analyzed the mechanism of αC polymer formation and tested our hypothesis that self-association of the αC-domains occurs through the interaction between their N-terminal subdomains and may include β-hairpin swapping. Our binding experiments performed by size-exclusion chromatography and optical trap-based force spectroscopy revealed that the αC-domains self-associate exclusively through their N-terminal subdomains, while their C-terminal subdomains were found to interact with the αC-connectors that tether the αC-domains to the bulk of the molecule. This interaction should reinforce the structure of αC polymers and provide the proper orientation of their reactive residues for efficient cross-linking by factor XIIIa. Molecular modeling of self-association of the N-terminal subdomains confirmed that the hypothesized β-hairpin swapping does not impose any steric hindrance. To "freeze" the conformation of the N-terminal subdomain and prevent the hypothesized β-hairpin swapping, we introduced by site-directed mutagenesis an extra disulfide bond between two β-hairpins of the bovine Aα406-483 fragment corresponding to this subdomain. The experiments performed by circular dichroism revealed that Aα406-483 mutant containing Lys429Cys/Thr463Cys mutations preserved its β-sheet structure. However, in contrast to wild-type Aα406-483, this mutant had lower tendency for oligomerization, and its structure was not stabilized upon oligomerization, in agreement with the above hypothesis. On the basis of the results obtained and our previous findings, we propose a model of fibrin αC polymer structure and molecular mechanism of assembly.