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Abstract Recent studies have shifted the spotlight from adult disease to gametogenesis and embryo developmental events, and these are greatly affected by various environmental chemicals, such as drugs, metabolites, pollutants, and others. Growing research has highlighted the critical importance of identifying and understanding the roles of chemicals in reproductive biology. However, the functions and mechanisms of chemicals in reproductive processes remain incomplete. We developed a comprehensive database called the Reproductive Chemical Database (RCDB) ( https://yu.life.sjtu.edu.cn/ChenLab/RCDB ) to facilitate research on chemicals in reproductive biology. This resource is founded on rigorous manual literature extraction and precise protein target prediction methodologies. This database focuses on the delineation of chemicals associated with phenotypes, diseases, or endpoints intricately associated with four important reproductive processes: female and male gamete generation, fertilization, and embryo development in human and mouse. The RCDB encompasses 93 sub-GO processes, and it revealed 1447 intricate chemical–biological process interactions. To date, the RCDB has meticulously cataloged and annotated 830 distinct chemicals, while also predicting 614 target proteins from a selection of 3800 potential candidates. Additionally, the RCDB offers an online predictive tool that empowers researchers to ascertain whether specific chemicals play discernible functional roles in these reproductive processes. The RCDB is an exhaustive, cross-platform, manually curated database, which provides a user-friendly interface to search, browse, and use reproductive processes modulators and their comprehensive related information. The RCDB will help researchers to understand the whole reproductive process and related diseases and it has the potential to promote reproduction research in the pharmacological and pathophysiological areas. Graphical Abstract

In this study, we report for the first time a new gabbro diorite, forming in an intraplate extensional environment during the Early Devonian, in the Shimian of the western margin of the Yangtze Block. Zircon SIMS U–Pb dating indicates that the gabbro diorite formed at ca. 406 Ma. This gabbro diorite belongs to low-K tholeiite and has enriched in Rb, Ba, K, Pb, negative anomalies of Nb, Ta, depletion of flat LREE patterns, negative εHf(t) (−1.1 to −11.7), and high δ18O (8.83 ± 0.21‰) ratios, showing that the magma formed by partial melting of N-MROB-type mantle and indicating that the magma underwent crustal contamination during emplacement. The magma is formed in the extensional environment, which provides evidence for the extensional environment in the western margin of the Yangtze. The geochemical characteristics and emplacement time of the rocks are similar to those in the early Paleozoic post-orogenic mafic rocks in South China, suggesting that the gabbro diorite may be influenced by the early Paleozoic orogeny in South China.

The Zhesang gold deposit is a Carlin-type gold deposit located in the “Golden Triangle” structural belt of Yunnan-Guizhou-Guangxi. Pyrite and arsenopyrite are important gold-bearing minerals of the deposit. In order to understand the typomorphic characteristics of pyrite and arsenopyrite and the enrichment mechanism of gold, morphological and structural features of these sulphides and the occurrence state of gold were investigated by using reflected light microscope and electron probes (EPMA). The results show that pyrites are characterized mainly by medium-fine broken pentagonal-dodecahedron crystal forms. Most of the arsenopyrites display needle and columnar crystal forms. Compositionally, pyrite is depleted in both Fe and S, whereas arsenopyrite is As-depleted but S-enriched. The w(Co)/w(Ni) of pyrite is between 1.731 and 3.215 (i.e., within the range of ratio of the hydrothermal pyrite, which is between 1 and 5). The crystal forms and compositional features of pyrite and arsenopyrite indicate that the Zhesang gold deposit is typical of medium-low temperature hydrothermal origin. The positive correlation between As and Au and the Au/As (mol%) ratio are lower than the solubility limit of Au in arsenic pyrite. Combined with previous studies, our data suggest that gold is likely to exist in the form of Au+ complex. Arsenic pyrite and arsenopyrite condensed Au in unsaturated ore-forming hydrothermal fluids relative to Au0 through surface voids and/or defects and the As content, forming a Zhesang Carlin-type gold deposit. Based on the age data of the Mesozoic polymetallic deposits in South China, it can be concluded that the age of the Zhesang gold deposit (215.3 ± 1.9 Ma), is consistent with the age of the main body of the Mesozoic polymetallic deposits (230–200 Ma) in South China Plate and the age of the granites (230–200 Ma) related to tungsten-tin mineralization in the Cathaysian Block. Therefore, we believe that the Zhesang gold deposit is the product of the subduction regime of the Paleo-Tethys Ocean in the Middle-Late Triassic.

Abstract Objective The purpose of this study was to explore the effect of cigarette smoke component (CSC) exposure on serum lipid levels in rats and the underlying molecular mechanism. Methods Male SPF-grade SD rats were randomly divided into a control group and a CSC exposure group, with the CSC group being exposed to CSC for 6 weeks. RT–PCR and Western blotting methods were used to detect lipid metabolism gene expression in rats, and 16S RNA gene sequencing was used to detect the gut microbiota in the rat cecum. Rat serum exosomes were prepared and identified, and the interaction of exosomal miR-291a-3p and miR-126a-5p with AMPK and CYP7A1 was detected by a dual luciferase reporter gene assay (DLRG). Results Serum indicators, including cholesterol levels and trimethylamine oxide (TMAO) content, were significantly affected in the CSC exposure group compared with the control group (P

BackgroundDisordered gut microbiota (GM) structure and function may contribute to osteoporosis (OP). This study explores how traditional Chinese medicine (TCM) intervention affects the structure and function of the GM in patients with OP.MethodIn a 3-month clinical study, 43 patients were randomly divided into two groups receiving conventional treatment and combined TCM (Yigu decoction, YGD) treatment. The correlation between the intestinal flora and its metabolites was analyzed using 16S rDNA and untargeted metabolomics and the combination of the two.ResultsAfter three months of treatment, patients in the treatment group had better bone mineral density (BMD) than those in the control group (P < 0.05). Patients in the treatment group had obvious abundance changes in GM microbes, such as Bacteroides, Escherichia-Shigella, Faecalibacterium, Megamonas, Blautia, Klebsiella, Romboutsia, Akkermansia, and Prevotella_9. The functional changes observed in the GM mainly involved changes in metabolic function, genetic information processing and cellular processes. The metabolites for which major changes were observed were capsazepine, Phe-Tyr, dichlorprop, D-pyroglutamic acid and tamsulosin. These metabolites may act through metabolic pathways, the citrate cycle (TCA cycle) and beta alanine metabolism. Combined analysis showed that the main acting metabolites were dichlorprop, capsazepine, D-pyroglutamic acid and tamsulosin.ConclusionThis study showed that TCM influenced the structure and function of the GM in patients with OP, which may be one mechanism by which TCM promotes the rehabilitation of patients with OP through the GM.

Two new species belonging to the varius group of Coccophagus, C. breviclavulus sp. nov. and C. perlucidus sp. nov., are described from Xishuangbanna Rainforest (China, Yunnan). Coccophagus anchoroides (Huang) and C. yunnana Wang, Huang & Polaszek are recorded. A tentative key to world species of this group is provided. Partial nuclear ribosomal 28S-D2 of these four species and other six species were sequenced and subjected to a phylogenetic analysis. Phylogeny of C. varius group is discussed.

The genus Eutrichosomella Girault is recorded for the first time from China (Yunnan Province), and Eutrichosomella yunnanensis sp. nov. (♀, ♂) is described and illustrated. A distribution map of this genus is presented.

With the worldwide epidemics of hyperuricemia and associated gout, the diseases with purine metabolic disorders have become a serious threat to human public health. Accumulating evidence has shown that they have been linked to increased consumption of fructose in humans, we hereby made a timely review on the roles of fructose intake and the gut microbiota in regulating purine metabolism, together with the potential mechanisms by which excessive fructose intake contributes to hyperuricemia and gout. To this end, we focus on the understanding of the interaction between a fructose-rich diet and the gut microbiota in hyperuricemia and gout to seek for safe, cheap, and side-effect-free clinical interventions. Furthermore, fructose intake recommendations for hyperuricemia and gout patients, as well as the variety of probiotics and prebiotics with uric acid-lowering effects targeting the intestinal tract are also summarized to provide reference and guidance for the further research.

Two new species of Paraphytis Compere, P. bannaensis sp. nov. and P. pseudovittatus sp. nov., are described from the Xishuangbanna Rainforest (Southwest China). A key to species from China based on females is provided.

Intrinsically disordered proteins (IDPs) account for more than 50% of the human proteome and are closely associated with tumors, cardiovascular diseases, and neurodegeneration, which have no fixed three-dimensional structure under physiological conditions. Due to the characteristic of conformational diversity, conventional experimental methods of structural biology, such as NMR, X-ray diffraction, and CryoEM, are unable to capture conformational ensembles. Molecular dynamics (MD) simulation can sample the dynamic conformations at the atomic level, which has become an effective method for studying the structure and function of IDPs. However, the high computational cost prevents MD simulations from being widely used for IDPs conformational sampling. In recent years, significant progress has been made in artificial intelligence, which makes it possible to solve the conformational reconstruction problem of IDP with fewer computational resources. Here, based on short MD simulations of different IDPs systems, we use variational autoencoders (VAEs) to achieve the generative reconstruction of IDPs structures and include a wider range of sampled conformations from longer simulations. Compared with the generative autoencoder (AEs), VAEs add an inference layer between the encoder and decoder in the latent space, which can cover the conformational landscape of IDPs more comprehensively and achieve the effect of enhanced sampling. Through experimental verification, the Cα RMSD between VAE-generated and MD simulation sampling conformations in the 5 IDPs test systems was significantly lower than that of AE. The Spearman correlation coefficient on the structure was higher than that of AE. VAE can also achieve excellent performance regarding structured proteins. In summary, VAEs can be used to effectively sample protein structures.

Phosphorylation plays a key role in plant biology, such as the accumulation of plant cells to form the observed proteome. Statistical analysis found that many phosphorylation sites are located in disordered regions. However, current force fields are mainly trained for structural proteins, which might not have the capacity to perfectly capture the dynamic conformation of the phosphorylated proteins. Therefore, we evaluated the performance of ff03CMAP, a balanced force field between structural and disordered proteins, for the sampling of the phosphorylated proteins. The test results of 11 different phosphorylated systems, including dipeptides, disordered proteins, folded proteins, and their complex, indicate that the ff03CMAP force field can better sample the conformations of phosphorylation sites for disordered proteins and disordered regions than ff03. For the solvent model, the results strongly suggest that the ff03CMAP force field with the TIP4PD water model is the best combination for the conformer sampling. Additional tests of CHARMM36m and FB18 force fields on two phosphorylated systems suggest that the overall performance of ff03CMAP is similar to that of FB18 and better than that of CHARMM36m. These results can help other researchers to choose suitable force field and solvent models to investigate the dynamic properties of phosphorylation proteins.

Chlorothricin (CHL) belongs to a spirotetronate antibiotic family produced by Streptomyces antibioticus that inhibits pyruvate carboxylase and malate dehydrogenase. For the biosynthesis of CHL, ChlB3 plays a crucial role by introducing the 6-methylsalicylic acid (6MSA) moiety to ChlB2, an acyl carrier protein (ACP). However, the structural insight and catalytic mechanism of ChlB3 was unclear. In the current study, the crystal structure of ChlB3 was solved at 3.1 Å-resolution and a catalytic mechanism was proposed on the basis of conserved residues of structurally related enzymes. ChlB3 is a dimer having the same active sites as CerJ (a structural homologous enzyme) and uses a KSIII-like fold to work as an acyltransferase. The relaxed substrate specificity of ChlB3 was defined by its catalytic efficiencies (kcat/Km) for non-ACP tethered synthetic substrates such as 6MSA-SNAC, acetyl-SNAC, and cyclohexonyl-SNAC. ChlB3 successfully detached the 6MSA moiety from 6MSA-SNAC substrate and this hydrolytic activity demonstrated that ChlB3 has the potential to catalyze non-ACP tethered substrates. Structural comparison indicated that ChlB3 belongs to FabH family and showed 0.6–2.5 Å root mean square deviation (RMSD) with structural homologous enzymes. Molecular docking and dynamics simulations were implemented to understand substrate active site and structural behavior such as the open and closed conformation of the ChlB3 protein. The resultant catalytic and substrate recognition mechanism suggested that ChlB3 has the potential to use non-native substrates and minimize the labor of expressing ACP protein. This versatile acyltransferase activity may pave the way for manufacturing CHL variants and may help to hydrolyze several thioester-based compounds.

Abstract As major environment factors, drought or high salinity affect crop growth, development and yield. Transgenic approach is an effective way to improve abiotic stress tolerance of crops. In this study, we comparatively analyzed gene structures, genome location, and the evolution of syntaxin proteins containing late embryogenesis abundant (LEA2) domain. GmSYP24 was identified as a dehydration-responsive gene. Our study showed that the GmSYP24 protein was located on the cell membrane. The overexpression of GmSYP24 (GmSYP24ox) in soybean and heteroexpression of GmSYP24 (GmSYP24hx) in Arabidopsis exhibited insensitivity to osmotic/drought and high salinity. However, wild type soybean, Arabidopsis, and the mutant of GmSYP24 homologous gene of Arabidopsis were sensitive to the stresses. Under the abiotic stresses, transgenic soybean plants had greater water content and higher activities of POD, SOD compared with non-transgenic controls. And the leaf stomatal density and opening were reduced in transgenic Arabidopsis. The sensitivity to ABA was decreased during seed germination of GmSYP24ox and GmSYP24hx. GmSYP24hx induced up-regulation of ABA-responsive genes. GmSYP24ox alters the expression of some aquaporins under osmotic/drought, salt, or ABA treatment. These results demonstrated that GmSYP24 played an important role in osmotic/drought or salt tolerance in ABA signal pathway.

Two new (1–2) and three known (3–5) sorbicillinoids were isolated from the deep-sea-derived fungus Penicillium allii-sativi MCCC 3A00580. Compounds 1 and 2, named sorbicatechols C and D, were two new hybrid dihydrosorbillinoids. Their structures were established mainly by spectroscopic analyses and electronic circular dichroism (ECD) calculations. All five isolates were tested for antiproliferative activities against four tumor cell lines of MCF-7, HT-29, HuH-7, and LNCap. Compounds 2 and 5 inhibited HT-29 cells in a good dose-dependent manner. Mechanism investigation uncovered that they could significantly induce cell cycle G2-M phase arresting by increasing the protein levels of p-H3 and cyclin B1.

The spontaneous depurination of genomic DNA occurs frequently and generates apurinic/pyrimidinic (AP) site damage that is mutagenic or lethal to cells. Error-prone DNA polymerases are specifically responsible for the translesion synthesis (TLS) of specific DNA damage, such as AP site damage, generally with relatively low fidelity. The Y-family DNA polymerases are the main error-prone DNA polymerases, and they employ three mechanisms to perform TLS, including template-skipping, dNTP-stabilized misalignment, and misincorporation-misalignment. The bypass mechanism of the dinB homolog (Dbh), an archaeal Y-family DNA polymerase from Sulfolobus acidocaldarius, is unclear and needs to be confirmed. In this study, we show that the Dbh primarily uses template skipping accompanied by dNTP-stabilized misalignment to bypass AP site analogs, and the incorporation of the first nucleotide across the AP site is the most difficult. Furthermore, based on the reported crystal structures, we confirmed that three conserved residues (Y249, R333, and I295) in the little finger (LF) domain and residue K78 in the palm subdomain of the catalytic core domain are very important for TLS. These results deepen our understanding of how archaeal Y-family DNA polymerases deal with intracellular AP site damage and provide a biochemical basis for elucidating the intracellular function of these polymerases.

(±)-Hypersines A–C (1–3), the three pairs of enantiomerically pure monoterpenoid polyprenylated acylphloroglucinols with an unprecedented 6/6/5/4 fused ring system, were isolated from Hypericum elodeoides. Their structures, including absolute configurations, were elucidated by comprehensive spectroscopic data, single-crystal X-ray diffraction, and quantum chemical calculations. The plausible, biosynthetic pathway of 1–3 was proposed. Moreover, the bioactivity evaluation indicated that 1a might be a novel DNA damage response inhibitor, and could enhance MCF-7 cell sensitivity to the anticancer agent, camptothecin.

Two new (1, 2) and one known (3) meroterpenoids were isolated from the deep-sea-derived fungus Penicillium allii-sativi. The relative structures of new compounds were determined on the basis of an extensive analysis of the NMR and MS data, and the absolute configurations were established by ECD calculations. Andrastone A (1) is a rare andrastin bearing an unusual cyclopentan-1,3-dione. It shows a selectively antiproliferative effect against HepG2 tumor cells with an IC50 value of 7.8 μM. Mechanism study showed that apoptosis via Caspase and RXRα pathways are responsible for the inhibitory effect.

Five new ergostanes, penicisteroids D−H (1−5), were isolated from the liquid culture of the deep-sea-derived fungus Penicillium granulatum MCCC 3A00475, along with 27 known compounds. The structures of the new steroids were established mainly on the basis of extensive analysis of 1D and 2D NMR as well as HRESIMS data. Moreover, the absolute configurations of 1 were confirmed unambiguously by the single-crystal X-ray crystallography. Compounds 2 and 4–7 showed moderate antiproliferative effects selectively against 12 different cancer cell lines with IC50 values of around 5 μM. Compounds 2 and 6, potent RXRα binders with Kd values of 13.8 and 12.9 μM, respectively, could induce apoptosis by a Retinoid X Receptor (RXR)-α-dependent mechanism by regulating RXRα transcriptional expression and promoting the poly-ADP-ribose polymerase (PARP) cleavage. Moreover, they could inhibit proliferation by cell cycle arrest at the G0/G1 phase.

Objective To investigate the clinical characteristics, classification, surgical approach, complication and prognosis of cervical dumbbell-shaped tumors. Methods Twenty-six consecutive cases with cervical dumbbell-shaped tumors were retrospectively studied. According to tumor location by imaging examination, all tumors were divided into 3 types. Type Ⅰ (17 cases) was mostly intravertebral and foraminal. Surgery through posterior approach was performed and internal fixation was operated in 8 cases. Type Ⅱ (4 cases) was mostly paravertebral and foraminal. Surgery through the anterolateral approach was performed without internal fixation. Type Ⅲ (5 cases) was equalization of intravertebral and paravertebral, and underwent surgery through combined posterior-anterolateral approach and internal fixation was performed in all of those cases. If the unilateral facet joint was destroyed, internal fixation was necessary. Lateral mass screw internal fixation and transpedicular screw fixation supplemented by fusion with autologous iliac bone graft were used to maintain cervical spinal stability. Results Among 26 patients there were 19 schwannomas, 4 neurofibromas, 2 gangliocytoma and 1 spinal meningioma. Total and subtotal tumor resection was achieved in 23 and 3 patients respectively. Among them 50% (13/26) of the cases were used internal fixation including 8 TypeⅠand 5 Type Ⅲ patients. The follow-up period was from 7 to 62 months, and mean time was 30 months. Four cases (15.38%) were found local tumor recurrence. Two cases suffered with surgical infection and cerebrospinal fluid leakage. There was no spinal cord injury and spinal deformity. Conclusion In order to increase the total resection rate and decrease recurrence rate, surgical approach should be selected according to the imaging classification of tumors. Stability reconstruction is absolutely necessary for the patients with facet joint destroyed. doi: 10.3969/j.issn.1672-6731.2013.11.006Video: http://www.cjcnn.org/index.php/cjcnn/pages/view/v13n11a6

Objective To investigate the surgical treatment and clinical results for the primary basilar invagination (BI) with atlantoaxial dislocation (AAD). Methods A retrospective study was performed. The study included 89 patients who had primary BI and AAD were surgically treated in our hospital from January 2008 to December 2011. There were 28 males and 61 females, aged between 10 and 69 years (mean 45.42 years). All patients were treated by the same 3⁃step surgical method. The first step, reduction of the AAD was performed by homemade odontoid repositor intraoperatively through posterior approach; the second step, AO stainless steel plates were fixed between the occiput and C2, 3 lateral mass screws; and the third step, occipitocervical fusion were completed by autologous iliac crest graft. Operation effect was recorded during follow ⁃up period. Results Follow ⁃up period was 6 to 48 months. Clinical symptoms were improved in 82.93% patients after the surgery. Japanse Orthopaedic Association (JOA) score increased from preoperative (8.80 ± 1.36) points to postoperative (15.35 ± 1.47) points (t = 17.225, P = 0.001). In general, satisfactory decompression and bony fusion were shown on postoperative radiological examinations for all patients. Compared with pretreatment data, the postoperative imaging measurement showed that the mean data of atlanto⁃dental interval (ADI, 9.22 mm vs 3.72 mm) and vertical dimension from the top of odontoid process to Chamberlain line (10.41 mm vs 3.23 mm) were all reduced, and the cervicomedullary angle (130° vs 150°) and space available of spinal cord (SAC, 11.13 mm vs 15.54 mm) were all improved. Conclusion The one⁃off surgery of posterior reduction technique and fixing between occiput and C2, 3 lateral mass screws is a safe, easy, and effective treatment for patients with primary BI and AAD. However, the final operative efficacy remains to be studied in long⁃term follow⁃up trial. DOI：10.3969/j.issn.1672⁃6731.2012.04.013

Objective To explore the optimal moment of surgical management for the anterior circulation aneurysms, and to summarize the experience of microsurgical treatment for the anterior circulation aneurysms. Methods The clinical data of 210 patients with anterior circulation aneurysms were retrospectively analyzed. In 183 patients with subarachnoid hemorrhage, there were 82 cases of grade Ⅰ -Ⅱ, 77 cases of grade Ⅲ, 21 cases of grade Ⅳ, 3 cases of grade Ⅴ according to Hunt⁃Hess grade. Results 199 patients underwent aneurysm clipping and 11 patients were underwent aneurysm wrapping. According to Glasgow Outcome Scale (GOS) classification criteria for the prognosis, 97 cases (46.19% ) cured, 80 cases (38.09%) had slight disability, 24 cases (11.43%) showed severe disability, and 9 cases (4.29%) died. Conclusion In the microsurgery for anterior circulation aneurysms, the patients should be early operated according to the individual clinical signs. The judgment and skills in surgery for the subtypes of anterior circulation aneurysms are the key points to cure intracranial aneurysms successfully. DOI：10.3969/j.issn.1672⁃6731.2012.01.012

A novel furocoumarin derivative named oxyalloimperatorin (1), together with seventeen furocoumarins 2–18 were isolated from the radix of Angelica dahurica. The chemical structure of new metabolite was characterized by analysis of IR, NMR, and HR-ESI-MS spectroscopic data. Among the isolated compounds, 13, 16, and 18 (each at 20 μM) could significantly promote the gene transcriptional function of nuclear receptor RXRα. While 7–9, 13, 14, and the new structure 1 (each at 20 μM) showed significant reduction in RXRα gene transcriptional activities induced by 9-cis-retinoid acid. The findings indicated that these furocoumarin skeleton derivatives might hold beneficial effects on many intractable diseases, such as cancer and metabolic diseases, due to their potential activities on regulating the transcriptional activation function of RXRα.

Two new steroidal saponins (1 and 2) were isolated from the dried bulbs of Allium macrostemon Bunge. Their structures were elucidated by the spectral data as 26-O-β-D-glucopyranosyl-5α-furost-25 (27)-ene-3β, 12β, 22, 26-tetraol-3-O-β-D-glucopyranosyl (1→2) [β-D-glucopyranosyl (1→3)]-β-D-glucopyranosyl (1→4)-β-D-galactopyranoside (1) and 26-O-β-D-glucopyranosyl-5β-furost-20 (22)-25 (27)-dien-3β, 12β, 26-triol-3-O-β-D-glucopyranosyl (1→2)-β-D-galactopyranoside (2), respectively. Their cytotoxic activities on several cancer cell lines (MCF-7, NCI-H460, SF-268 and HepG2) were tested. 1 showed special cytotoxity on SF-268, while 2 showed cytotoxity on NCI-H460 and SF-268 cell lines, respectively.

Enzyme promiscuity is a prerequisite for fast divergent evolution of biocatalysts. A phosphotriesterase-like lactonase (PLL) from Geobacillus kaustophilus HTA426 (GkaP) exhibits main lactonase and promiscuous phosphotriesterase activities. To understand its catalytic and evolutionary mechanisms, we investigated a "hot spot" in the active site by saturation mutagenesis as well as X-ray crystallographic analyses. We found that position 99 in the active site was involved in substrate discrimination. One mutant, Y99L, exhibited 11-fold improvement over wild-type in reactivity (kcat/Km) toward the phosphotriesterase substrate ethyl-paraoxon, but showed 15-fold decrease toward the lactonase substrate δ-decanolactone, resulting in a 157-fold inversion of the substrate specificity. Structural analysis of Y99L revealed that the mutation causes a ∼6.6 Å outward shift of adjacent loop 7, which may cause increased flexibility of the active site and facilitate accommodation and/or catalysis of organophosphate substrate. This study provides for the PLL family an example of how the evolutionary route from promiscuity to specificity can derive from very few mutations, which promotes alteration in the conformational adjustment of the active site loops, in turn draws the capacity of substrate binding and activity.

The transactivation domain (TAD) of tumor suppressor p53 can bind with the nuclear coactivator binding domain (NCBD) of cyclic-AMP response element binding protein (CBP) and activate transcription. NMR experiments demonstrate that both apo-NCBD and TAD are intrinsic disordered and bound NCBD/TAD undergoes a transition to well folded. The recognition mechanism between intrinsic disordered proteins is still hotly debated. Molecular dynamics (MD) simulations in explicit solvent are used to study the recognition mechanism between intrinsic disordered TAD and NCBD. The average RMSD values between bound and corresponding apo states and Kolmogorov-Smirnov P test analysis indicate that TAD and NCBD may follow an induced fit mechanism. Quantitative analysis indicates there is also a global conformational selection. In summary, the recognition of TAD and NCBD might obey a local induced fit and global conformational selection. These conclusions are further supported by high-temperature unbinding kinetics and room temperature landscape analysis. These methods can be used to study the recognition mechanism of other intrinsic disordered proteins.

Amyloid fibrils are found in many fatal neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, type II diabetes, and prion disease. The VEALYL short peptide from insulin has been confirmed to aggregate amyloid-like fibrils. However, the aggregation mechanism of amyloid fibril is poorly understood. Here, we utilized molecular dynamics simulation to analyse the stability of VEALYL hexamer. The statistical results indicate that hydrophobic residues play key roles in stabilizing VEALYL hexamer. Single point and two linkage mutants confirmed that Val1, Leu4, and Tyr5 of VEALYL are key residues. The consistency of the results for the VEALYL oligomer suggests that the intermediate states might be trimer (3-0) and pentamer(3-2). These results can help us to obtain an insight into the aggregation mechanism of amyloid fibril. These methods can be used to study the stability of amyloid fibril from other short peptides.

MicroRNAs are endogenous 23-25 nt RNAs that play important gene-regulatory roles in animals and plants. Recently, miR369-3 was found to upregulate translation of TNFα mRNA in quiescent (G0) mammalian cell lines. Knock down and immunofluorescence experiments suggest that microRNA-protein complexes (with FXR1 and AGO2) are necessary for the translation upregulation. However the molecular mechanism of microRNA translation activation is poorly understood. In this study we constructed the microRNA-mRNA-AGO2-FXR1 quadruple complex by bioinformatics and molecular modeling, followed with all atom molecular dynamics simulations in explicit solvent to investigate the interaction mechanisms for the complex. A combined analysis of experimental and computational data suggests that AGO2-FXR1 complex relocalize microRNA:mRNA duplex to polysomes in G0. The two strands of dsRNA are then separated upon binding of AGO2 and FXR1. Finally, polysomes may improve the translation efficiency of mRNA. The mutation research confirms the stability of microRNA-mRNA-FXR1 and illustrates importance of key residue of Ile304. This possible mechanism can shed more light on the microRNA-dependent upregulation of translation.

BACKGROUND:Kinase-inducible domain (KID) as transcriptional activator can stimulate target gene expression in signal transduction by associating with KID interacting domain (KIX). NMR spectra suggest that apo-KID is an unstructured protein. After post-translational modification by phosphorylation, KID undergoes a transition from disordered to well folded protein upon binding to KIX. However, the mechanism of folding coupled to binding is poorly understood. METHODOLOGY:To get an insight into the mechanism, we have performed ten trajectories of explicit-solvent molecular dynamics (MD) for both bound and apo phosphorylated KID (pKID). Ten MD simulations are sufficient to capture the average properties in the protein folding and unfolding. CONCLUSIONS:Room-temperature MD simulations suggest that pKID becomes more rigid and stable upon the KIX-binding. Kinetic analysis of high-temperature MD simulations shows that bound pKID and apo-pKID unfold via a three-state and a two-state process, respectively. Both kinetics and free energy landscape analyses indicate that bound pKID folds in the order of KIX access, initiation of pKID tertiary folding, folding of helix alpha(B), folding of helix alpha(A), completion of pKID tertiary folding, and finalization of pKID-KIX binding. Our data show that the folding pathways of apo-pKID are different from the bound state: the foldings of helices alpha(A) and alpha(B) are swapped. Here we also show that Asn139, Asp140 and Leu141 with large Phi-values are key residues in the folding of bound pKID. Our results are in good agreement with NMR experimental observations and provide significant insight into the general mechanisms of binding induced protein folding and other conformational adjustment in post-translational modification.