Water quality drives the reconfiguration of riverine planktonic microbial food webs.

The food web is a cycle of matter and energy within river ecosystems. River environmental changes resulting from human activities are increasingly threatening the composition and diversity of global aquatic organisms and the multi-trophic networks. How multiple environmental factors influence food web patterns among multi-trophic microbial communities in rivers remains largely unknown. Using water quality evaluation and meta-omics techniques, we investigated the composition, structure and interaction characteristics, and drivers of food webs of microorganisms (archaea, bacteria, fungi, protists, metazoa, viridiplantae and viruses) at multiple trophic levels in different water quality environments (Classes II, III, and IV). First, water quality deterioration led to significant changes in the composition of the microbial community at multiple trophic levels, which were represented by the enrichment of Euryarchaeota in the archaeal community, the increase of r-strategists in the bacterial community, and the increase of the proportion of predators in the protist community. Second, deteriorating water quality resulted in a significant reduction in the dissimilarity of community structure (homogenization of community structure in Class III and IV waters). Of the symbiotic, parasitic, and predatory networks, the community networks in Class II water all showed the most stable symbiotic, parasitic, and predatory correlations (higher levels of modularity in the networks). In Class III and IV waters, nutrient inputs have led to increased reciprocal symbiosis and decreased competition between communities, which may have the risk of a positive feedback loop driving a system collapse. Finally, inputs of phosphorus and organic matter could be the main drivers of changes in the planktonic microbial food web in the Fen River. Overall, the results indicated the potential ecological risks of exogenous nutrient inputs, which were important for aquatic ecosystem conservation.

Structural insights into the catalysis and substrate specificity of cyanobacterial aspartate racemase McyF.

L-amino acids represent the most common amino acid form, most notably as protein residues, whereas D-amino acids, despite their rare occurrence, play significant roles in many biological processes. Amino acid racemases are enzymes that catalyze the interconversion of L- and/or D-amino acids. McyF is a pyridoxal 5'-phosphate (PLP) independent amino acid racemase that produces the substrate D-aspartate for the biosynthesis of microcystin in the cyanobacterium Microcystis aeruginosa PCC7806. Here we report the crystal structures of McyF in complex with citrate, L-Asp and D-Asp at 2.35, 2.63 and 2.80 Å, respectively. Structural analyses indicate that McyF and homologs possess highly conserved residues involved in substrate binding and catalysis. In addition, residues Cys87 and Cys195 were clearly assigned to the key catalytic residues of "two bases" that deprotonate D-Asp and L-Asp in a reaction independent of PLP. Further site-directed mutagenesis combined with enzymatic assays revealed that Glu197 also participates in the catalytic reaction. In addition, activity assays proved that McyF could also catalyze the interconversion of L-MeAsp between D-MeAsp, the precursor of another microcystin isoform. These findings provide structural insights into the catalytic mechanism of aspartate racemase and microcystin biosynthesis.

Structural insights into SraP-mediated Staphylococcus aureus adhesion to host cells.

Staphylococcus aureus, a Gram-positive bacterium causes a number of devastating human diseases, such as infective endocarditis, osteomyelitis, septic arthritis and sepsis. S. aureus SraP, a surface-exposed serine-rich repeat glycoprotein (SRRP), is required for the pathogenesis of human infective endocarditis via its ligand-binding region (BR) adhering to human platelets. It remains unclear how SraP interacts with human host. Here we report the 2.05 Å crystal structure of the BR of SraP, revealing an extended rod-like architecture of four discrete modules. The N-terminal legume lectin-like module specifically binds to N-acetylneuraminic acid. The second module adopts a ß-grasp fold similar to Ig-binding proteins, whereas the last two tandem repetitive modules resemble eukaryotic cadherins but differ in calcium coordination pattern. Under the conditions tested, small-angle X-ray scattering and molecular dynamic simulation indicated that the three C-terminal modules function as a relatively rigid stem to extend the N-terminal lectin module outwards. Structure-guided mutagenesis analyses, in addition to a recently identified trisaccharide ligand of SraP, enabled us to elucidate that SraP binding to sialylated receptors promotes S. aureus adhesion to and invasion into host epithelial cells. Our findings have thus provided novel structural and functional insights into the SraP-mediated host-pathogen interaction of S. aureus.

Structure of a novel O-linked N-acetyl-D-glucosamine (O-GlcNAc) transferase, GtfA, reveals insights into the glycosylation of pneumococcal serine-rich repeat adhesins.

Protein glycosylation catalyzed by the O-GlcNAc transferase (OGT) plays a critical role in various biological processes. In Streptococcus pneumoniae, the core enzyme GtfA and co-activator GtfB form an OGT complex to glycosylate the serine-rich repeat (SRR) of adhesin PsrP (pneumococcal serine-rich repeat protein), which is involved in the infection and pathogenesis. Here we report the 2.0 Å crystal structure of GtfA, revealing a ß-meander add-on domain beyond the catalytic domain. It represents a novel add-on domain, which is distinct from the all-α-tetratricopeptide repeats in the only two structure-known OGTs. Structural analyses combined with binding assays indicate that this add-on domain contributes to forming an active GtfA-GtfB complex and recognizing the acceptor protein. In addition, the in vitro glycosylation system enables us to map the O-linkages to the serine residues within the first SRR of PsrP. These findings suggest that fusion with an add-on domain might be a universal mechanism for diverse OGTs that recognize varying acceptor proteins/peptides.

Structure of the adenylation-peptidyl carrier protein didomain of the Microcystis aeruginosa microcystin synthetase McyG.

Microcystins, which are the most common cause of hepatotoxicity associated with cyanobacterial water blooms, are assembled in vivo on a large multienzyme complex via a mixed nonribosomal peptide synthetase/polyketide synthetase (NRPS/PKS). The biosynthesis of microcystin in Microcystis aeruginosa PCC 7806 starts with the enzyme McyG, which contains an adenylation-peptidyl carrier protein (A-PCP) didomain for loading the starter unit to assemble the side chain of an Adda residue. However, the catalytic mechanism remains unclear. Here, the 2.45 Šresolution crystal structure of the McyG A-PCP didomain complexed with the catalytic intermediate L-phenylalanyl-adenylate (L-Phe-AMP) is reported. Each asymmetric unit contains two protein molecules, one of which consists of the A-PCP didomain and the other of which comprises only the A domain. Structural analyses suggest that Val227 is likely to be critical for the selection of hydrophobic substrates. Moreover, two distinct interfaces demonstrating variable crosstalk between the PCP domain and the A domain were observed. A catalytic cycle for the adenylation and peptide transfer of the A-PCP didomain is proposed.

Structural insights into the substrate specificity of a 6-phospho-ß-glucosidase BglA-2 from Streptococcus pneumoniae TIGR4.

The 6-phospho-ß-glucosidase BglA-2 (EC 3.2.1.86) from glycoside hydrolase family 1 (GH-1) catalyzes the hydrolysis of ß-1,4-linked cellobiose 6-phosphate (cellobiose-6'P) to yield glucose and glucose 6-phosphate. Both reaction products are further metabolized by the energy-generating glycolytic pathway. Here, we present the first crystal structures of the apo and complex forms of BglA-2 with thiocellobiose-6'P (a non-metabolizable analog of cellobiose-6'P) at 2.0 and 2.4 Å resolution, respectively. Similar to other GH-1 enzymes, the overall structure of BglA-2 from Streptococcus pneumoniae adopts a typical (ß/α)8 TIM-barrel, with the active site located at the center of the convex surface of the ß-barrel. Structural analyses, in combination with enzymatic data obtained from site-directed mutant proteins, suggest that three aromatic residues, Tyr(126), Tyr(303), and Trp(338), at subsite +1 of BglA-2 determine substrate specificity with respect to 1,4-linked 6-phospho-ß-glucosides. Moreover, three additional residues, Ser(424), Lys(430), and Tyr(432) of BglA-2, were found to play important roles in the hydrolytic selectivity toward phosphorylated rather than non-phosphorylated compounds. Comparative structural analysis suggests that a tryptophan versus a methionine/alanine residue at subsite -1 may contribute to the catalytic and substrate selectivity with respect to structurally similar 6-phospho-ß-galactosidases and 6-phospho-ß-glucosidases assigned to the GH-1 family.

Structure and catalytic mechanism of yeast 4-amino-4-deoxychorismate lyase.

Saccharomyces cerevisiae Abz2 is a pyridoxal 5'-phosphate (PLP)-dependent lyase that converts 4-amino-4-deoxychorismate (ADC) to para-aminobenzoate and pyruvate. To investigate the catalytic mechanism, we determined the 1.9 Å resolution crystal structure of Abz2 complexed with PLP, representing the first eukaryotic ADC lyase structure. Unlike Escherichia coli ADC lyase, whose dimerization is critical to the formation of the active site, the overall structure of Abz2 displays as a monomer of two domains. At the interdomain cleft, a molecule of cofactor PLP forms a Schiff base with residue Lys-251. Computational simulations defined a basic clamp to orientate the substrate ADC in a proper pose, which was validated by site-directed mutageneses combined with enzymatic activity assays. Altogether, we propose a putative catalytic mechanism of a unique class of monomeric ADC lyases led by yeast Abz2.

Crystal structures and catalytic mechanism of the C-methyltransferase Coq5 provide insights into a key step of the yeast coenzyme Q synthesis pathway.

Saccharomyces cerevisiae Coq5 is an S-adenosyl methionine (SAM)-dependent methyltransferase (SAM-MTase) that catalyzes the only C-methylation step in the coenzyme Q (CoQ) biosynthesis pathway, in which 2-methoxy-6-polyprenyl-1,4-benzoquinone (DDMQH2) is converted to 2-methoxy-5-methyl-6-polyprenyl-1,4-benzoquinone (DMQH2). Crystal structures of Coq5 were determined in the apo form (Coq5-apo) at 2.2 Šresolution and in the SAM-bound form (Coq5-SAM) at 2.4 Šresolution, representing the first pair of structures for the yeast CoQ biosynthetic enzymes. Coq5 displays a typical class I SAM-MTase structure with two minor variations beyond the core domain, both of which are considered to participate in dimerization and/or substrate recognition. Slight conformational changes at the active-site pocket were observed upon binding of SAM. Structure-based computational simulation using an analogue of DDMQH2 enabled us to identify the binding pocket and entrance tunnel of the substrate. Multiple-sequence alignment showed that the residues contributing to the dimeric interface and the SAM- and DDMQH2-binding sites are highly conserved in Coq5 and homologues from diverse species. A putative catalytic mechanism of Coq5 was proposed in which Arg201 acts as a general base to initiate catalysis with the help of a water molecule.

Optimal format design in green tourism supply chain in the presence of a dominant online travel agency.

Due to a large consumer base owned by a dominant online travel agency(OTA), green hotels widely cooperate with the OTA to sell their products. This study develops a green tourism supply chain model involving an OTA and a green hotel. The study aims to provide valuable insights for the OTA and hotel in optimizing their decision-making process when they come to reach an agreement on cooperation formats among price-only, cost sharing, and revenue sharing. The findings indicate that cost sharing and revenue sharing formats are more effective than price-only format in incentivizing the hotel to improve its environmental efforts. Moreover, these two formats result in higher wholesale prices determined by the hotel compared to the wholesale price set in price-only format. This study also recommends the revenue sharing format as an optimal format, which could lead to a win-win outcome for both the OTA and hotel, achieving profit maximization under specific conditions. Overall, by effectively implementing cooperation formats, the OTA and hotel can work together to promote green tourism supply chain performances.

Combining spectral and texture feature of UAV image with plant height to improve LAI estimation of winter wheat at jointing stage.

Introduction: Leaf area index (LAI) is a critical physiological and biochemical parameter that profoundly affects vegetation growth. Accurately estimating the LAI for winter wheat during jointing stage is particularly important for monitoring wheat growth status and optimizing variable fertilization decisions. Recently, unmanned aerial vehicle (UAV) data and machine/depth learning methods are widely used in crop growth parameter estimation. In traditional methods, vegetation indices (VI) and texture are usually to estimate LAI. Plant Height (PH) unlike them, contains information about the vertical structure of plants, which should be consider. Methods: Taking Xixingdian Township, Cangzhou City, Hebei Province, China as the research area in this paper, and four machine learning algorithms, namely, support vector machine(SVM), back propagation neural network (BPNN), random forest (RF), extreme gradient boosting (XGBoost), and two deep learning algorithms, namely, convolutional neural network (CNN) and long short-term memory neural network (LSTM), were applied to estimate LAI of winter wheat at jointing stage by integrating the spectral and texture features as well as the plant height information from UAV multispectral images. Initially, Digital Surface Model (DSM) and Digital Orthophoto Map (DOM) were generated. Subsequently, the PH, VI and texture features were extracted, and the texture indices (TI) was further constructed. The measured LAI on the ground were collected for the same period and calculated its Pearson correlation coefficient with PH, VI and TI to pick the feature variables with high correlation. The VI, TI, PH and fusion were considered as the independent features, and the sample set partitioning based on joint x-y distance (SPXY) method was used to divide the calibration set and validation set of samples. Results: The ability of different inputs and algorithms to estimate winter wheat LAI were evaluated. The results showed that (1) The addition of PH as a feature variable significantly improved the accuracy of the LAI estimation, indicating that wheat plant height played a vital role as a supplementary parameter for LAI inversion modeling based on traditional indices; (2) The combination of texture features, including normalized difference texture indices (NDTI), difference texture indices (DTI), and ratio texture indices (RTI), substantially improved the correlation between texture features and LAI; Furthermore, multi-feature combinations of VI, TI, and PH exhibited superior capability in estimating LAI for winter wheat; (3) Six regression algorithms have achieved high accuracy in estimating LAI, among which the XGBoost algorithm estimated winter wheat LAI with the highest overall accuracy and best results, achieving the highest R2 (R2 = 0.88), the lowest RMSE (RMSE=0.69), and an RPD greater than 2 (RPD=2.54). Discussion: This study provided compelling evidence that utilizing XGBoost and integrating spectral, texture, and plant height information extracted from UAV data can accurately monitor LAI during the jointing stage of winter wheat. The research results will provide a new perspective for accurate monitoring of crop parameters through remote sensing.

Structural insights into HetR-PatS interaction involved in cyanobacterial pattern formation.

The one-dimensional pattern of heterocyst in the model cyanobacterium Anabaena sp. PCC 7120 is coordinated by the transcription factor HetR and PatS peptide. Here we report the complex structures of HetR binding to DNA, and its hood domain (HetRHood) binding to a PatS-derived hexapeptide (PatS6) at 2.80 and 2.10 Å, respectively. The intertwined HetR dimer possesses a couple of novel HTH motifs, each of which consists of two canonical α-helices in the DNA-binding domain and an auxiliary α-helix from the flap domain of the neighboring subunit. Two PatS6 peptides bind to the lateral clefts of HetRHood, and trigger significant conformational changes of the flap domain, resulting in dissociation of the auxiliary α-helix and eventually release of HetR from the DNA major grove. These findings provide the structural insights into a prokaryotic example of Turing model.