The formation of Jupiter's diluted core by a giant impact.

The Juno mission1 has provided an accurate determination of Jupiter's gravitational field2, which has been used to obtain information about the planet's composition and internal structure. Several models of Jupiter's structure that fit the probe's data suggest that the planet has a diluted core, with a total heavy-element mass ranging from ten to a few tens of Earth masses (about 5 to 15 per cent of the Jovian mass), and that  heavy elements (elements other than hydrogen and helium) are distributed within a region extending to nearly half of Jupiter's radius3,4. Planet-formation models indicate that most heavy elements are accreted during the early stages of a planet's formation to create a relatively compact core5-7 and that almost no solids are accreted during subsequent runaway gas accretion8-10. Jupiter's diluted core, combined with its possible high heavy-element enrichment, thus challenges standard planet-formation theory. A possible explanation is erosion of the initially compact heavy-element core, but the efficiency of such erosion is uncertain and depends on both the immiscibility of heavy materials in metallic hydrogen and on convective mixing as the planet evolves11,12. Another mechanism that can explain this structure is planetesimal enrichment and vaporization13-15 during the formation process, although relevant models typically cannot produce an extended diluted core. Here we show that a sufficiently energetic head-on collision (giant impact) between a large planetary embryo and the proto-Jupiter could have shattered its primordial compact core and mixed the heavy elements with the inner envelope. Models of such a scenario lead to an internal structure that is consistent with a diluted core, persisting over billions of years. We suggest that collisions were common in the young Solar system and that a similar event may have also occurred for Saturn, contributing to the structural differences between Jupiter and Saturn16-18.

Evaluation of parametrial infiltration in patients with IB-IIB cervical cancer by a radiomics model integrating features from tumoral and peritumoral regions in 18 F-fluorodeoxy glucose positron emission tomography/MR images.

Parametrial infiltration (PMI) is an essential factor in staging and planning treatment of cervical cancer. The purpose of this study was to develop a radiomics model for accessing PMI in patients with IB-IIB cervical cancer using features from 18 F-fluorodeoxy glucose (18 F-FDG) positron emission tomography (PET)/MR images. In this retrospective study, 66 patients with International Federation of Gynecology and Obstetrics stage IB-IIB cervical cancer (22 with PMI and 44 without PMI) who underwent 18 F-FDG PET/MRI were divided into a training dataset (n = 46) and a testing dataset (n = 20). Features were extracted from both the tumoral and peritumoral regions in 18 F-FDG PET/MR images. Single-modality and multimodality radiomics models were developed with random forest to predict PMI. The performance of the models was evaluated with F1 score, accuracy, and area under the curve (AUC). The Kappa test was used to observe the differences between PMI evaluated by radiomics-based models and pathological results. The intraclass correlation coefficient for features extracted from each region of interest (ROI) was measured. Three-fold crossvalidation was conducted to confirm the diagnostic ability of the features. The radiomics models developed by features from the tumoral region in T2 -weighted images (F1 score = 0.400, accuracy = 0.700, AUC = 0.708, Kappa = 0.211, p = 0.329) and the peritumoral region in PET images (F1 score = 0.533, accuracy = 0.650, AUC = 0.714, Kappa = 0.271, p = 0.202) achieved the best performances in the testing dataset among the four single-ROI radiomics models. The combined model using features from the tumoral region in T2 -weighted images and the peritumoral region in PET images achieved the best performance (F1 score = 0.727, accuracy = 0.850, AUC = 0.774, Kappa = 0.625, p < 0.05). The results suggest that 18 F-FDG PET/MRI can provide complementary information regarding cervical cancer. The radiomics-based method integrating features from the tumoral and peritumoral regions in 18 F-FDG PET/MR images gave a superior performance for evaluating PMI.

Opportunity cost determines free-operant action initiation latency and predicts apathy.

BACKGROUND: Apathy, a disabling and poorly understood neuropsychiatric symptom, is characterised by impaired self-initiated behaviour. It has been hypothesised that the opportunity cost of time (OCT) may be a key computational variable linking self-initiated behaviour with motivational status. OCT represents the amount of reward which is foregone per second if no action is taken. Using a novel behavioural task and computational modelling, we investigated the relationship between OCT, self-initiation and apathy. We predicted that higher OCT would engender shorter action latencies, and that individuals with greater sensitivity to OCT would have higher behavioural apathy. METHODS: We modulated the OCT in a novel task called the 'Fisherman Game', Participants freely chose when to self-initiate actions to either collect rewards, or on occasion, to complete non-rewarding actions. We measured the relationship between action latencies, OCT and apathy for each participant across two independent non-clinical studies, one under laboratory conditions (n = 21) and one online (n = 90). 'Average-reward' reinforcement learning was used to model our data. We replicated our findings across both studies. RESULTS: We show that the latency of self-initiation is driven by changes in the OCT. Furthermore, we demonstrate, for the first time, that participants with higher apathy showed greater sensitivity to changes in OCT in younger adults. Our model shows that apathetic individuals experienced greatest change in subjective OCT during our task as a consequence of being more sensitive to rewards. CONCLUSIONS: Our results suggest that OCT is an important variable for determining free-operant action initiation and understanding apathy.

New insights into the dihydro-mureidomycin biosynthesis controlled by two unusual proteins in Streptomyces roseosporus.

BACKGROUND: Uridyl peptide compounds are renowned as a subclass of nucleoside antibiotics for their highly specific antibacterial activity against Gram-negative bacteria and the unique target of action. We previously activated the biosynthetic gene cluster of a uridyl peptide antibiotic, mureidomycin, in Streptomyces roseosporus NRRL 15998 by introducing an exogenous positive regulator gene ssaA, and the generated strain was designated as Sr-hA. This study aims to further explore mureidomycin analogs from Sr-hA as well as the collaborative roles of two wide-spread genes, SSGG-02980 and SSGG-03002 encoding putative nuclease/phosphatase and oxidoreductase respectively, in mureidomycin diversification. RESULTS: In order to understand how SSGG-02980 and SSGG-03002 contribute to mureidomycin biosynthesis, the gene disruption mutants and complementary strains were constructed. Mass spectrometry analyses revealed that two series of pairwise mureidomycin analogs were synthesized in Sr-hA with a two-dalton difference in molecular weight for each pair. By disruption of SSGG-03002, only mureidomycins with lower molecular weight (MRDs, 1-6) could be specifically accumulated in the mutant (∆03002-hA), whereas the other series of products with molecular weight plus 2 Da (rMRDs, 1'-6') became dominant in SSGG-02980 disruption mutant (∆02980-hA). Further comprehensive NMR analyses were performed to elucidate the structures, and three MRDs (3, 4, 5) with unsaturated double bond at C5-C6 of uracil group were characterized from ∆03002-hA. In contrast, the paired rMRDs analogs (3', 4', 5') from ∆SSGG-02980 corresponding to 3, 4 and 5 were shown to contain a single bond at this position. The results verified that SSGG-03002 participates in the reduction of uracil ring, whereas SSGG-02980 antagonizes the effect of SSGG-03002, which has been rarely recognized for a phosphatase. CONCLUSIONS: Overall, this study revealed the key roles of two wide-spread families of enzymes in Streptomyces. Of them, oxidoreductase, SSGG-03002, is involved in dihydro-mureidomycin biosynthesis of S. roseosporus, whereas nuclease/phosphatase, SSGG-02980, has an adverse effect on SSGG-03002. This kind of unusual regulation model between nuclease/phosphatase and oxidoreductase is unprecedented, providing new insights into the biosynthesis of mureidomycins in Streptomyces. The findings would be of significance for structural diversification of more uridyl peptide antibiotics against Gram-negative bacteria.

The Role of eIF5A1 in LPS-Induced Neuronal Remodeling of the Nucleus Accumbens in the Depression.

BACKGROUND: The pathogenesis of depression is complex, with the brain's reward system likely to play an important role. The nucleus accumbens (NAc) is a key region in the brain that integrates reward signals. Lipopolysaccharides (LPS) can induce depressive-like behaviors and enhance neuroplasticity in NAc, but the underlying mechanism is still unknown. We previously found that eukaryotic translation initiation factor A1 (eIF5A1) acts as a ribosome-binding protein to regulate protein translation and to promote neuroplasticity. METHODS: In the present study, LPS was administered intraperitoneally to rats and the expression and cellular location of eIF5A1 was then investigated by RT-PCR, Western blotting and immunofluorescence. Subsequently, a neuron-specific lentivirus was used to regulate eIF5A1 expression in vivo and in vitro. Neuroplasticity was then examined by Golgi staining and by measurement of neuronal processes. Finally, proteomic analysis was used to identify proteins regulated by eIF5A1. RESULTS: The results showed that eIF5A1 expression was significantly increased in the NAc neurons of LPS rats. Following the knockdown of eIF5A1 in NAc neurons, the LPS-induced increases in neuronal arbors and spine density were significantly attenuated. Depression-like behaviors were also reduced. Neurite outgrowth of NAc neurons in vitro also increased or decreased in parallel with the increase or decrease in eIF5A1 expression, respectively. The proteomic results showed that eIF5A1 regulates the expression of many neuroplasticity-related proteins in neurons. CONCLUSIONS: These results confirm that eIF5A1 is involved in LPS-induced depression-like behavior by increasing neuroplasticity in the NAc. Our study also suggests the brain's reward system may play an important role in the pathogenesis of depression.

Stress conditions induced circRNAs profile of extracellular vesicles in brain microvascular endothelial cells.

Cerebral ischemia causes hypoxic injury and inflammation, and brain microvascular endothelial cells (BMVECs) dysfunction is an initial stage of blood-brain barrier disruption. Endothelial cells secrete extracellular vesicles (EVs) that are involved in intercellular signal transduction. EVs contain a variety of RNAs, proteins, and metabolites. Circular RNA (circRNA) is a member of the non-coding RNA. The expression profile and potential function of circRNAs in BMVECs are unknown. Here, human BMVECs have undergone hypoxia or TNF-α induction, and the changes in circRNAs were measured by RNA sequencing. A total of 70 circRNAs showed differential expression, including 43 previously unrecorded circRNAs and 27 recorded circRNAs. Since astrocyte end-feet encircle endothelial cells, they are considered the main targets of the EVs from BMVEC. The miRNA sequence data and bioinformatics were used to predict the circRNA-miRNA-mRNA networks in astrocytes. The gene ontology (GO) analysis showed the main downstream targets of circRNAs are DNA transcription regulation and protein kinase-related signaling pathways. These results suggest that altering circRNAs may be a potential therapeutic target for cerebral ischemia induced hypoxic injury and inflammation.

Typical Fragment Kinetic Energy Assessment Based on Acoustic Emission Technology.

Fragment kinetic energy is an important parameter to characterize the damage power of fragments. In this study, an acoustic emission technology-based method to evaluate fragment kinetic energy is proposed. The dynamic response of the fragment impacting an aluminum alloy target plate and the relationship between the initial kinetic energy of the fragment impact and the acoustic emission waveform were theoretically evaluated; the numerical simulation of typical spherical fragments (8 mm diameter) penetrating the aluminum alloy target plate was performed, the wavelet energy of the acoustic emission signal was obtained using wavelet packet theory, and a mathematical model of wavelet energy and fragment kinetic energy was constructed. A fragment kinetic energy test system was established, and a fragment penetration test was performed. The analysis showed that the wavelet energy mathematical models and the fragment kinetic energy exhibited favorable consistency, and the measurement errors of the three experiments were 3%, 3.7%, and 3%. This demonstrates the effectiveness of the typical acoustic emission fragment kinetic energy test methods proposed in this study and establishes a new method for the direct measurement of fragment kinetic energy.

Soil actinobacteria tend to have neutral interactions with other co-occurring microorganisms, especially under oligotrophic conditions.

Actinobacteria produce a variety of secondary metabolites that can influence the survival or behaviour of other organisms. The understanding of the ecological roles of actinobacteria has significantly improved in the past decades, but a systematic insight into the interactions between actinobacteria and other microbes in nature is warranted. Here, we studied the pairwise effects of actinobacteria on other microbes isolated from red soils under different nutritional conditions. We found that neutral effects dominated the interactions, accounting for 68.1% of the interactions in eutrophic conditions and for a significantly higher proportion (86.2%) in oligotrophic conditions. High nutrient levels boosted active metabolism of actinobacteria and generally made them more aggressive, supporting the stress gradient hypothesis. The secondary metabolites produced by actinobacteria played a pivotal role in interference competition with other microbes, of which the role of desferrioxamine siderophores could not be ignored. Niche overlap seemed to be another cause of competition, notably under oligotrophic conditions. Moreover, the large-scale phylogeny had a much greater impact on the interaction than the location origin of the microbes. These results provide an understanding of the coexistence of actinobacteria with other microbes in nature and suggest neutrality as a key mechanism for maintaining microbial diversity in soils.

LsrR, the effector of AI-2 quorum sensing, is vital for the H2O2 stress response in mammary pathogenic Escherichia coli.

Mammary pathogenic Escherichia coli (MPEC) is an important causative agent of mastitis in dairy cows that results in reduced milk quality and production, and is responsible for severe economic losses in the dairy industry worldwide. Oxidative stress, as an imbalance between reactive oxygen species (ROS) and antioxidants, is a stress factor that is common in most bacterial habitats. The presence of ROS can damage cellular sites, including iron-sulfur clusters, cysteine and methionine protein residues, and DNA, and may cause bacterial cell death. Previous studies have reported that Autoinducer 2 (AI-2) can regulate E. coli antibiotic resistance and pathogenicity by mediating the intracellular receptor protein LsrR. This study explored the regulatory mechanism of LsrR on the H2O2 stress response in MPEC, showing that the transcript levels of lsrR significantly decreased under H2O2 stress conditions. The survival cell count of lsrR mutant XW10/pSTV28 was increased about 3080-fold when compared with that of the wild-type WT/pSTV28 in the presence of H2O2 and overexpression of lsrR (XW10/pUClsrR) resulted in a decrease in bacterial survival rates under these conditions. The ß-galactosidase reporter assays showed that mutation of lsrR led to a remarkable increase in expression of the promoters of ahpCF, katG and oxyR, while lsrR-overexpressing significantly reduced the expression of ahpCF and katG. The electrophoretic mobility shift assays confirmed that LsrR could directly bind to the promoter regions of ahpCF and katG. These results revealed the important role played by LsrR in the oxidative stress response of MPEC.

Broad and dynamic neurochemical alterations in the brain of alcoholic rats.

Ethanol is the active ingredient in alcoholic beverages. As ethanol consumption increases from zero to very high, it is still unknown which metabolites are present at different times and which are essential to normal functioning. In this article, we used an intermittent-access 20% ethanol drinking paradigm to make Wistar male rats voluntarily drink large amounts of ethanol for 10, 20, 30, and 50 days, respectively. A hydrogen-1 nuclear magnetic resonance approach was used to investigate the time-dependent neurochemical metabolites spectra in the hippocampus, striatum, nucleus accumbens and prefrontal cortex. Multivariate pattern recognition techniques were used to analyze the hydrogen-1 nuclear magnetic resonance spectra data. Metabolic profiling was obtained, differentiating the ethanol-treated and control rats. The ethanol-affected metabolites disrupted processes associated with neurotransmitters, oxidative stress, energy metabolism and amino acids. Together, our findings demonstrate broad, dynamic, and time-dependent endogenous metabolic alterations in rats treated with ethanol.

Route of administration of exogenous progesterone for luteal support does not significantly affect the serum concentration in assisted reproductive technology.

The objective of this research is to study the guiding role of serum progesterone level on exogenous luteal support protocols. In the retrospective study, a total of 537 infertile women undergoing IVF/ICSI were recruited. Serum samples were obtained for serum progesterone measurements. The results demonstrated that the progesterone levels of all women gradually decreased over the course of 7 days after ET. The progesterone level of the pregnant women reached a nadir on day 7 after ET and subsequently began to rise, while the progesterone level of the non-pregnant women continued to decrease. Even with different routes of administration of exogenous progesterone, the progesterone levels followed the same patterns. The serum progesterone level does not represent the adequacy of exogenous progesterone supplementation. Therefore, there is no need to measure serum progesterone levels frequently after embryo transfer or adjust the dose according to serum progesterone levels.

Progesterone promotes endothelial nitric oxide synthase expression through enhancing nuclear progesterone receptor-SP-1 formation.

Progesterone exerts antihypertensive actions partially by modulating endothelial nitric oxide synthase (eNOS) activity. Here, we aimed to investigate the effects and mechanisms of progesterone on eNOS expression. First, human umbilical vein endothelial cells (HUVECs) were exposed to progesterone and then the eNOS transcription factor specificity protein-1 (SP-1) and progesterone receptor (PRA/B) expression were assessed by Western blotting and qRT-PCR. The interaction between SP-1 and PRA/B was next determined through coimmunoprecipitation assay. The chromatin immunoprecipitation assay and luciferase assay were used to investigate the relationship of PRA/B, SP-1, and eNOS promoter. At last, rats were intraperitoneally injected with progesterone receptor antagonist RU-486, and then the expression of eNOS and vasodilation function in thoracic aorta and mesenteric artery were measured. The results showed that progesterone could increase eNOS expression in HUVECs. Further study showed that progesterone increased PRA-SP-1 complex formation and facilitated PRA/B and SP-1 binding to eNOS promoter. Mutating SP-1 or PR-binding motif on eNOS promoter abolished the effect of progesterone on eNOS gene transcription. We also observed that progesterone receptor antagonist RU-486 reduced eNOS expression and impaired vasodilation in rats. Those results suggest that progesterone modulates eNOS expression through promoting PRA-SP-1 complex formation, and progesterone antagonist attenuates eNOS expression, leading to the loss of vascular relaxation.NEW & NOTEWORTHY Progesterone directly upregulated endothelial nitric oxide synthase (eNOS) expression in human endothelial cells. Progesterone augmented eNOS promoter activity through a progesterone receptor A- and specificity protein-1-dependent manner. Antagonism of the progesterone receptor reduced eNOS expression and impaired vasodilation in rats.

Myocardial tissue and metabolism characterization in men with alcohol consumption by cardiovascular magnetic resonance and 11C-acetate PET/CT.

BACKGROUND: Chronic alcohol consumption initially leads to asymptomatic left ventricular dysfunction, but can result in myocardial impairment and heart failure if ongoing. This study sought to characterize myocardial tissues and oxidative metabolism in asymptomatic subjects with chronic alcohol consumption by quantitative cardiovascular magnetic resonance (CMR) and 11C-acetate positron emission tomography (PET)/computed tomography (CT). METHODS: Thirty-four male subjects (48.8 ± 9.1 years) with alcohol consumption > 28 g/day for > 10 years and 35 age-matched healthy male subjects (49.5 ± 9.7 years) underwent CMR and 11C-acetate PET/CT. Native and post T1 values and extracellular volume (ECV) from CMR and Kmono and K1 from PET imaging were measured. Quantitative measurements by CMR and PET imaging were compared between subjects with moderate to heavy alcohol consumption and healthy controls, and their correlations were also analyzed. RESULTS: Compared to healthy controls, subjects with alcohol consumption showed significantly shorter native T1 (1133 ± 65 ms vs. 1186 ± 31 ms, p < 0.001) and post T1 (477 ± 42 ms vs. 501 ± 38 ms, p = 0.008) values, greater ECV (28.2 ± 2.2% vs. 26.9 ± 1.3%, p = 0.003), marginally lower Kmono (57.6 ± 12.1 min- 1 × 10- 3 vs. 63.0 ± 11.7 min- 1 × 10- 3, p = 0.055), and similar K1 (0.82 ± 0.13 min- 1 vs. 0.83 ± 0.15 min- 1, p = 0.548) after adjusting for confounding factors. There were no significant differences in CMR measurements and K1 between subjects with heavy and moderate alcohol consumption (all p > 0.05). In contrast, subjects with heavy alcohol consumption showed significantly lower Kmono values compared to those with moderate alcohol consumption (52.9 ± 12.1 min- 1 × 10- 3 vs. 63.7 ± 9.2 min- 1 × 10- 3, p = 0.012). Strong and moderate correlations were found between K1 and ECV in healthy controls (r = 0.689, p = 0.013) and subjects with moderate alcohol consumption (r = 0.518, p = 0.048), respectively. CONCLUSION: Asymptomatic men with heavy alcohol consumption have detectable structural and metabolic changes in myocardium on CMR and 11C-acetate PET/CT. Compared with quantitative CMR, 11C-acetate PET/CT imaging may be more sensitive for detecting differences in myocardial damage among subjects with moderate to heavy alcohol consumption.

The two-component system, BasSR, is involved in the regulation of biofilm and virulence in avian pathogenic Escherichia coli.

Avian pathogenic Escherichia coli (APEC) is a subgroup of extra-intestinal pathogenic E. coli (ExPEC) strains that cause avian colibacillosis, resulting in significant economic losses to the poultry industry worldwide. It has been reported that a few two-component signal transduction systems (TCS) participate in the regulation of the virulence factors of APEC infection. In this study, a basSR-deficient mutant strain was constructed from its parent strain APECX40 (WT), and high-throughput sequencing (RNA-seq) was performed to analyse the transcriptional profile of WT and its mutant strain XY1. Results showed that the deletion of basSR down-regulated the transcript levels of a series of biofilm- and virulence-related genes. Results of biofilm formation assays and bird model experiments indicated that the deletion of basSR inhibited biofilm formation in vitro and decreased bacterial virulence and colonization in vivo. In addition, electrophoretic mobility shift assays confirmed that the BasR protein could bind to the promoter regions of several biofilm- and virulence-related genes, including ais, opgC and fepA. This study suggests that the BasSR TCS might be a global regulator in the pathogenesis of APEC infection. RESEARCH HIGHLIGHTS Transcriptional profiling showed that BasSR might be a global regulator in APEC. BasSR increases APEC pathogenicity in vivo. BasSR positively regulates biofilm- and the virulence-associated genes. BasSR can bind to the promoter regions of virulence-associated genes ais, opgC and fepA.

N-Glycosylation at Asn695 might suppress inducible nitric oxide synthase activity by disturbing electron transfer.

Inducible nitric oxide synthase (iNOS) plays critical roles in the inflammatory response and host defense. Previous research on iNOS regulation mainly focused on its gene expression level, and much less is known about the regulation of iNOS function by N-glycosylation. In this study, we report for the first time that iNOS is N-glycosylated in vitro and in vivo. Mass spectrometry studies identified Asn695 as an N-glycosylation site of murine iNOS. Mutating Asn695 to Gln695 yields an iNOS that exhibits greater enzyme activity. The essence of nitric oxide synthase catalytic reaction is electron transfer process, which involves a series of conformational changes, and the linker between the flavin mononucleotide-binding domain and the flavin adenine dinucleotide-binding domain plays vital roles in the conformational changes. Asn695 is part of the linker, so we speculated that attachment of N-glycan to the Asn695 residue might inhibit activity by disturbing electron transfer. Indeed, our NADPH consumption results demonstrated that N-glycosylated iNOS consumes NADPH more slowly. Taken together, our results indicate that iNOS is N-glycosylated at its Asn695 residue and N-glycosylation of Asn695 might suppress iNOS activity by disturbing electron transfer.

Crystal Structure of Aeromonas hydrophila Cytoplasmic 5'-Methylthioadenosine/S-Adenosylhomocysteine Nucleosidase.

5'-Methylthioadenosine/S-adenosyl-l-homocysteine (MTA/SAH) nucleosidase (MTAN) is an important enzyme in a number of critical biological processes. Mammals do not express MtaN, making this enzyme an attractive antibacterial drug target. In pathogen Aeromonas hydrophila, two MtnN subfamily genes (MtaN-1 and MtaN-2) play important roles in the periplasm and cytosol, respectively. We previously reported structural and functional analyses of MtaN-1, but little is known regarding MtaN-2 due to the lack of a crystal structure. Here, we determined the crystal structure of cytosolic A. hydrophila MtaN-2 in complex with adenine (ADE), which is a cleavage product of adenosine. AhMtaN-1 and AhMtaN-2 exhibit a high degree of similarity in the α-ß-α sandwich fold of the core structural motif. However, there is a structural difference in the nonconserved extended loop between ß7 and α3 that is associated with the channel depth of the substrate-binding pocket and dimerization. The ADE molecules in the substrate-binding pockets of AhMtaN-1 and AhMtaN-2 are stabilized with π-π stacking by Trp199 and Phe152, respectively, and the hydrophobic residues surrounding the ribose-binding sites differ. A structural comparison of AhMtaN-2 with other MtaN proteins showed that MtnN subfamily proteins exhibit a unique substrate-binding surface and dimerization interface.

Structural insight into the carboxylesterase BioH from Klebsiella pneumoniae.

The BioH carboxylesterase which is a typical α/ß-hydrolase enzyme involved in biotin synthetic pathway in most bacteria. BioH acts as a gatekeeper and blocks the further elongation of its substrate. In the pathogen Klebsiella pneumoniae, BioH plays a critical role in the biosynthesis of biotin. To better understand the molecular function of BioH, we determined the crystal structure of BioH from K. pneumoniae at 2.26 Šresolution using X-ray crystallography. The structure of KpBioH consists of an α-ß-α sandwich domain and a cap domain. B-factor analysis revealed that the α-ß-α sandwich domain is a rigid structure, while the loops in the cap domain shows the structural flexibility. The active site of KpBioH contains the catalytic triad (Ser82-Asp207-His235) on the interface of the α-ß-α sandwich domain, which is surrounded by the cap domain. Size exclusion chromatography shows that KpBioH prefers the monomeric state in solution, whereas two-fold symmetric dimeric formation of KpBioH was observed in the asymmetric unit, the conserved Cys31-based disulfide bonds can maintain the irreversible dimeric formation of KpBioH. Our study provides important structural insight for understanding the molecular mechanisms of KpBioH and its homologous proteins.

Crystal structure of the Siderophore-interacting protein SIP from Aeromonas hydrophila.

Siderophores acquire iron from hosts under iron-limiting conditions and play an essential role in the survival of microorganisms. Siderophore-interacting proteins (SIPs) from microbes release iron from the siderophore complex by reducing ferric iron to ferrous iron, but the molecular mechanism of iron reduction remains unclear. To better understand the molecular mechanism of SIPs, we herein report the crystal structure of Aeromonas hydrophila SIP (AhSIP) in complex with flavin adenine dinucleotide (FAD) as a cofactor. AhSIP consists of an N-terminal FAD binding domain and a C-terminal NADH binding domain, which are connected by a linker region. AhSIP showed unique structural differences in the orientation of the cofactor binding lobes when compared with SIP homologs. This study identified a cluster of three basic residues (Lys48, His259 and Arg262) in AhSIP distributed around a potential substrate binding pocket. In addition, AhSIP, containing the NADH binding motif E(L)VL-X3-GE, belongs to the group I subfamily. Our results show the diverse cofactor and substrate binding sites of the SIP family.

Structural and functional analyses of the lipase CinB from Enterobacter asburiae.

Lipases are widely present in various plants, animals and microorganisms, constituting a large category of enzymes. They have the ability to catalyze the cleavage of ester bonds. The lipase CinB from Enterobacter asburiae (E. asburiae) is an acetyl esterase. The primary amino acid sequence suggests that the EaCinB protein belongs to the α/ß-hydrolase (ABH) superfamily of the esterase/lipase superfamily. However, its molecular functions have not yet been determined. Here, we report the crystal structure of E. asburiae CinB at a 1.45 Šresolution. EaCinB contains a signal peptide, cap domain and catalytic domain. The active site of EaCinB contains the catalytic triad (Ser180-His307-Asp277) on the catalytic domain. The oxyanion hole is composed of Gly106 and Gly107 within the conserved sequence motif HGGG (amino acid residues 106-109). The substrate is accessible between the α1 and α2 helices or the α1 helix and catalytic domain. Narrow substrate pockets are formed by the α2 helix of the cap domain. Site-directed mutagenesis showed that EaCinB-W208H exhibits a higher catalytic ability than EaCinB-WT by approximately nine times. Our results provide insight into the molecular function of EaCinB.

Structural insight of the 5-(Hydroxyethyl)-methylthiazole kinase ThiM involving vitamin B1 biosynthetic pathway from the Klebsiella pneumoniae.

Thiamin pyrophosphate (TPP) is an essential co-factor in amino acid and carbohydrate metabolic pathways. The TPP-related vitamin B1 biosynthetic pathway is found in most bacterial, plant and lower eukaryotic processes; however, it is not present in humans. In bacterial thiamin synthesis and salvage pathways, the 5-(hydroxyethyl)-methylthiazole kinase (ThiM) is essential in the pathway forming TPP. Thus, ThiM is considered to be an attractive antibacterial drug target. Here, we determined the crystal structures of ThiM from pathogenic Klebsiella pneumoniae (KpThiM) and KpThiM in complex with its substrate 5-(hydroxyethyl)-4-methylthiazole (TZE). KpThiM, consisting of an α-ß-α domain, shows a pseudosymmetric trimeric formation. TZE molecules are located in the interface between the KpThiM subunits in the trimer and interact with Met49 and Cys200. Superimposition of the apo and TZE-complexed structures of KpThiM show that the side chains of the amino acids interacting with TZE and Mg2+ have a rigid configuration. Comparison of the ThiM structures shows that KpThiM could, in terms of sequence and configuration, be different from other ThiM proteins, which possess different amino acids that recognize TZE and Mg2+. The structures will provide new insight into the ThiM subfamily proteins for antibacterial drug development.