Cilia are slender, micrometer-long organelles present on the surface of eukaryotic cells. They function in signaling and locomotion and are constructed by intraflagellar transport (IFT). The assembly of IFT complexes into so-called IFT trains to initiate ciliary entry at the base of the cilium remains a matter of debate. Here, we use structural modeling to provide an architectural framework for how RabL2 is anchored at the ciliary base via CEP19 before being handed over to IFT trains for ciliary entry. Our models suggest that the N-terminal domain of CEP43 forms a homo-dimer to anchor at the subdistal appendages of cilia through a direct interaction with CEP350. A long linker region separates the N-terminal domain of CEP43 from the C-terminal domain, which captures CEP19 above the subdistal appendages and close to the distal appendages. Furthermore, we present a structural model for how RabL2-CEP19 associates with the IFT-B complex, providing insight into how RabL2 is handed over from CEP19 to the IFT complex. Interestingly, RabL2 association with the IFT-B complex appears to induce a significant conformational change in the IFT complex via a kink in the coiled-coils of the IFT81/74 proteins, which may prime the IFT machinery for entry into cilia.

A propeptide is removed from a precursor protein to generate its active or mature form. Propeptides play essential roles in protein folding, transportation, and activation and are present in about 2.3% of reviewed proteins in the UniProt database. They are often found in secreted or membrane-bound proteins including proteolytic enzymes, hormones, and toxins. We identified a variety of globular and nonglobular Pfam domains in protein sequences designated as propeptides, some of which form intramolecular interactions with other domains in the mature proteins. Propeptide-containing enzymes mostly function as proteases, as they are depleted in other enzyme classes such as hydrolases acting on DNA and RNA, isomerases, and lyases. We applied AlphaFold to generate structural models for over 7000 proteins with propeptides having no less than 20 residues. Analysis of residue contacts in these models revealed conformational changes for over 300 proteins before and after the cleavage of the propeptide. Examples of conformation change occur in several classes of proteolytic enzymes in the families of subtilisins, trypsins, aspartyl proteases, and thermolysin-like metalloproteases. In most of the observed cases, cleavage of the propeptide releases the constraints imposed by the covalent bond between the propeptide and the mature protein, and cleavage enables stronger interactions between the propeptide and the mature protein. These findings suggest that post-cleavage propeptides could play critical roles in regulating the activity of mature proteins.

In the current study, we utilized molecular modeling and simulation approaches to define putative potential molecular targets for Burdock Inulin, including inflammatory proteins such as iNOS, COX-2, TNF-alpha, IL-6, and IL-1β. Molecular docking results revealed potential interactions and good binding affinity for these targets; however, IL-1β, COX-2, and iNOS were identified as the best targets for Inulin. Molecular simulation-based stability assessment demonstrated that inulin could primarily target iNOS and may also supplementarily target COX-2 and IL-1β during DSS-induced colitis to reduce the role of these inflammatory mechanisms. Furthermore, residual flexibility, hydrogen bonding, and structural packing were reported with uniform trajectories, showing no significant perturbation throughout the simulation. The protein motions within the simulation trajectories were clustered using principal component analysis (PCA). The IL-1β-Inulin complex, approximately 70% of the total motion was attributed to the first three eigenvectors, while the remaining motion was contributed by the remaining eigenvectors. In contrast, for the COX2-Inulin complex, 75% of the total motion was attributed to the eigenvectors. Furthermore, in the iNOS-Inulin complex, the first three eigenvectors contributed to 60% of the total motion. Furthermore, the iNOS-Inulin complex contributed 60% to the total motion through the first three eigenvectors. To explore thermodynamically favorable changes upon mutation, motion mode analysis was carried out. The Free Energy Landscape (FEL) results demonstrated that the IL-1β-Inulin achieved a single conformation with the lowest energy, while COX2-Inulin and iNOS-Inulin exhibited two lowest-energy conformations each. IL-1β-Inulin and COX2-Inulin displayed total binding free energies of - 27.76 kcal/mol and - 37.78 kcal/mol, respectively, while iNOS-Inulin demonstrated the best binding free energy results at - 45.89 kcal/mol. This indicates a stronger pharmacological potential of iNOS than the other two complexes. Thus, further experiments are needed to use inulin to target iNOS and reduce DSS-induced colitis and other autoimmune diseases.

This study examined the effect of the human-functioning dimension on happiness among community-dwelling older adults (OAs) in Chile. Questionnaires were used for data collection from a sample of 785 OAs of both sexes attending healthcare institutions. Exploratory factor analysis was performed using parallel analysis and oblique rotation. Confirmatory factor analysis and structural equation modeling were conducted using the maximum likelihood and unweighted least squares methods. Goodness-of-fit analyses were performed by considering absolute and respective incremental fit indices. The relationships between the functioning and happiness factors were all significant at the 1% level, indicating that functioning impacts happiness. The ratios of the variances between both constructs were identical to those of the covariances, indicating consistency between the models, with similarities and equalities in the estimation of their parameters. The modeling confirms a direct relationship between activities of daily living functioning and happiness. Given that a lack of functioning significantly affects OAs\' happiness and quality of life, this relationship is consistent with the available theory. These findings may contribute to the formulation of social and health policies regarding OAs in Chile and other Latin American countries.

Drosophila has been used as an animal model to study pathogenic mechanism of neurological disorders. Thymidylate kinase (TMPK) is an essential enzyme in dTTP synthesis catalyzing the phosphorylation of dTMP to dTDP. Loss of function mutations in the DTYMK gene, coding for TMPK, cause severe microcephaly in human patients. In this study, Drosophila melanogaster TMPK (DmTMPK) was cloned, expressed, purified and characterized. Unlike human TMPK, DmTMPK phosphorylated not only dTMP and dUMP but also dGMP and dIMP although with low efficiency. ATP and dATP are the most efficient phosphate donor but at higher concentration (>1 mM) ATP inhibited DmTMPK activity. Sequence and structural model analysis explain why DmTMPK could phosphorylate purine nucleoside monophosphates. This study has laid a solid foundation for future study of TMPK function in Drosophila.

Coat proteins (CP) of the potato virus A virions (PVA) contain partially disordered N-terminal domains, which are necessary for performing vital functions of the virus. Comparative analysis of the structures of coat proteins (CPs) in the intact PVA virions and in the virus particles lacking N-terminal 32 amino acids (PVAΔ32) was carried out in this work based on the tritium planigraphy data. Using atomic-resolution structure of the potato virus Y potyvirus (PVY) protein, which is a homolog of the CP PVA, the available CP surfaces in the PVY virion were calculated and the areas of intersubunit/interhelix contacts were determined. For this purpose, the approach of Lee and Richards [Lee, B., and Richards, F. M. (1971) J. Mol. Biol., 55, 379-400] was used. Comparison of incorporation profiles of the tritium label in the intact and trypsin-degraded PVAΔ32 revealed position of the ΔN-peptide shielding the surface domain (a.a. 66-73, 141-146) and the interhelix zone (a.a. 161-175) of the PVA CP. Presence of the channels/cavities was found in the virion, which turned out to be partially permeable to tritium atoms. Upon removal of the ΔN-peptide, decrease in the label incorporation within the virion (a.a. 184-200) was also observed, indicating possible structural transition leading to the virion compactization. Based on the obtained data, we can conclude that part of the surface ΔN-peptide is inserted between the coils of the virion helix thus increasing the helix pitch and providing greater flexibility of the virion, which is important for intercellular transport of the viruses in the plants.

Ion channels play key roles in human physiology and are important targets in drug discovery. The atomic-scale structures of ion channels provide invaluable insights into a fundamental understanding of the molecular mechanisms of channel gating and modulation. Recent breakthroughs in deep learning-based computational methods, such as AlphaFold, RoseTTAFold, and ESMFold have transformed research in protein structure prediction and design. We review the application of AlphaFold, RoseTTAFold, and ESMFold to structural modeling of ion channels using representative voltage-gated ion channels, including human voltage-gated sodium (NaV) channel - NaV1.8, human voltage-gated calcium (CaV) channel - CaV1.1, and human voltage-gated potassium (KV) channel - KV1.3. We compared AlphaFold, RoseTTAFold, and ESMFold structural models of NaV1.8, CaV1.1, and KV1.3 with corresponding cryo-EM structures to assess details of their similarities and differences. Our findings shed light on the strengths and limitations of the current state-of-the-art deep learning-based computational methods for modeling ion channel structures, offering valuable insights to guide their future applications for ion channel research.

Estrogens play critical roles in embryonic development, gonadal sex differentiation, behavior, and reproduction in vertebrates and in several human cancers. Estrogens are synthesized from testosterone and androstenedione by the endoplasmic reticulum membrane-bound P450 aromatase/cytochrome P450 oxidoreductase complex (CYP19/CPR). Here, we report the characterization of novel mammalian CYP19 isoforms encoded by CYP19 gene copies. These CYP19 isoforms are all defined by a combination of mutations in the N-terminal transmembrane helix (E42K, D43N) and in helix C of the catalytic domain (P146T, F147Y). The mutant CYP19 isoforms show increased androgen conversion due to the KN transmembrane helix. In addition, the TY substitutions in helix C result in a substrate preference for androstenedione. Our structural models suggest that CYP19 mutants may interact differently with the membrane (affecting substrate uptake) and with CPR (affecting electron transfer), providing structural clues for the catalytic differences.

The structural modeling of peptides can be a useful aid in the discovery of new drugs and a deeper understanding of the molecular mechanisms of life. Here we present a novel multiscale protocol for the structure prediction of linear and cyclic peptides. The protocol combines two main stages: coarse-grained simulations using the CABS-flex standalone package and an all-atom reconstruction-optimization process using the Modeller program. We evaluated the protocol on a set of linear peptides and two sets of cyclic peptides, with cyclization through the backbone and disulfide bonds. A comparison with other state-of-the-art tools (APPTEST, PEP-FOLD, ESMFold and AlphaFold implementation in ColabFold) shows that for most cases, AlphaFold offers the highest resolution. However, CABS-flex is competitive, particularly when it comes to short linear peptides. As demonstrated, the protocol performance can be further improved by combination with the residue-residue contact prediction method or more efficient scoring. The protocol is included in the CABS-flex standalone package along with online documentation to aid users in predicting the structure of peptides and mini-proteins.

Stenotrophomonas maltophilia was discovered as a soil bacterium associated with the rhizosphere. Later, S. maltophilia was found to be a multidrug-resistant hospital-associated pathogen. Lytic bacteriophages are prospective antimicrobials; therefore, there is a need for the isolation and characterization of new Stenotrophomonas phages. The phage StenM_174 was isolated from litter at a poultry farm using a clinical strain of S. maltophilia as the host. StenM_174 reproduced in a wide range of clinical and environmental strains of Stenotrophomonas, mainly S. maltophilia, and it had a podovirus morphotype. The length of the genomic sequence of StenM_174 was 42,956 bp, and it contained 52 putative genes. All genes were unidirectional, and 31 of them encoded proteins with predicted functions, while the remaining 21 were identified as hypothetical ones. Two tail spike proteins of StenM_174 were predicted using AlphaFold2 structural modeling. A comparative analysis of the genome shows that the Stenotrophomonas phage StenM_174, along with the phages Ponderosa, Pepon, Ptah, and TS-10, can be members of the new putative genus Ponderosavirus in the Autographiviridae family. In addition, the analyzed data suggest a new subfamily within this family.