Structural insights and functional characterization of a novel ß-glucosidase derived from Thermotoga profunda.

ß-Glucosidase is a crucial cellulase, as its activity determines the efficiency of cellulose hydrolysis into glucose. This study addresses the functional and structural characteristics of Thermotoga profunda ß-glucosidase (Tp-BGL). Tp-BGL exhibited a Km of 0.3798 mM for p-nitrophenyl-ß-d-glucopyranoside (pNPGlc) and 4.44 mM for cellobiose, with kcat/Km of 1211.16 and 4.18 s-1 mM-1, respectively. In addition, Tp-BGL showed significant pH adaptability and thermal stability, with a Tm of 85.7 °C and retaining >90 % of its activity after incubation at 80 °C for 90 min. The crystal structure of Tp-BGL was resolved at 1.95 Å resolution, and reveals a typical TIM barrel structure. Comparative structural analysis highlighted that the major distinction between Tp-BGL and the other glucosidases lies in their loop regions.

Machine Learning-Driven Discovery and Evaluation of Antimicrobial Peptides from Crassostrea gigas Mucus Proteome.

Marine antimicrobial peptides (AMPs) represent a promising source for combating infections, especially against antibiotic-resistant pathogens and traditionally challenging infections. However, traditional drug discovery methods face challenges such as time-consuming processes and high costs. Therefore, leveraging machine learning techniques to expedite the discovery of marine AMPs holds significant promise. Our study applies machine learning to develop marine AMPs, focusing on Crassostrea gigas mucus rich in antimicrobial components. We conducted proteome sequencing of C. gigas mucous proteins, used the iAMPCN model for peptide activity prediction, and evaluated the antimicrobial, hemolytic, and cytotoxic capabilities of six peptides. Proteomic analysis identified 4490 proteins, yielding about 43,000 peptides (8-50 amino acids). Peptide ranking based on length, hydrophobicity, and charge assessed antimicrobial potential, predicting 23 biological activities. Six peptides, distinguished by their high relative scores and promising biological activities, were chosen for bactericidal assay. Peptides P1 to P4 showed antimicrobial activity against E. coli, with P2 and P4 being particularly effective. All peptides inhibited S. aureus growth. P2 and P4 also exhibited significant anti-V. parahaemolyticus effects, while P1 and P3 were non-cytotoxic to HEK293T cells at detectable concentrations. Minimal hemolytic activity was observed for all peptides even at high concentrations. This study highlights the potent antimicrobial properties of naturally occurring oyster mucus peptides, emphasizing their low cytotoxicity and lack of hemolytic effects. Machine learning accurately predicted biological activity, showcasing its potential in peptide drug discovery.

Characterization of pollen tube development in distant hybridization of Chinese cork oak (Quercus variabilis L.).

MAIN CONCLUSION: This work mainly found that the stigma and style of Q. variabilis did not completely lose the specific recognition towards heterologous pollen, a fact which is different from previous studies. Quercus is the foundation species in the Northern Hemisphere, with extreme prevalence for interspecific hybridization. It is not yet entirely understood whether or how the pollen tube-female tissue interaction contributes to the "extensive hybridization" in oaks. Pollen storage conditions correlate with distant hybridization. We conducted hybridization experiments with Q. variabilis as female and Q. variabilis and Q. mongolica as male parents. And the differences in pollen tube (PT) development between intra- and distant interspecific hybridization were studied by fluorescence microscopy and scanning electron microscopy (SEM). Our results showed that -20 °C allowed pollen of both species to maintain some viability. Both Q. variabilis and Q. mongolica pollen germinated profusely on the stigmas. SEM results indicated that in the intraspecific hybridization, Q. variabilis pollen started to germinate at 6 h after pollination (hap), PTs elongated significantly at 12 hap, and entered the stigma at 24 hap. By contrast, Q. mongolica pollen germinated at 15 hap, and the PTs entered the stigma at 27 hap. By fluorescence microscopical studies it was observed that some PTs of Q. variabilis gathered at the style-joining at 96 hap, unlike the Q. mongolica which reached the style junction at 144 hap. The above results indicate that the abundant germination of heterologous pollen (HP) on the stigma and the "Feeble specificity recognition" of the stigma and transmitting tract to HP may create opportunities for the "extensive hybridization" of oaks. This work provides a sexual developmental reference for clarifying the causes of Quercus "extensive hybridization".

iTRAQ-based proteomic analysis reveals the underlying mechanism of postmortem tenderization of refrigerated porcine Longissimus thoracis et lumborum muscle.

The isobaric tags for relative and absolute quantitation (iTRAQ) technology was used for differential proteomic analysis of refrigerated porcine Longissimus thoracis et lumborum (LTL) muscle at different time points postmortem (45 min, 4 h, 8 h, 12 h, 24 h, 48 h, 72 h and 96 h) to mechanistically elucidate the postmortem tenderization. Compared with the proteins identified in porcine LTL muscle at 45 min postmortem (control), 862 proteins were significantly expressed at 4 h, 8 h, 12 h, 24 h, 48 h, 72 h and 96 h postmortem. Moreover, clustering and path analysis showed that the quality traits of porcine LTL muscle, including pH, shear force, myofibril fragmentation index, correlated significantly with 2, 6 and 6 differentially expressed proteins, respectively, with the lowest or highest expression at 8 h or 12 h postmortem. Overall, the tenderness of refrigerated porcine LTL muscle might be significantly affected by changes in quality traits at 8 h and 12 h postmortem.

Multi-Exposure Image Fusion Algorithm Based on Improved Weight Function.

High-dynamic-range (HDR) image has a wide range of applications, but its access is limited. Multi-exposure image fusion techniques have been widely concerned because they can obtain images similar to HDR images. In order to solve the detail loss of multi-exposure image fusion (MEF) in image reconstruction process, exposure moderate evaluation and relative brightness are used as joint weight functions. On the basis of the existing Laplacian pyramid fusion algorithm, the improved weight function can capture the more accurate image details, thereby making the fused image more detailed. In 20 sets of multi-exposure image sequences, six multi-exposure image fusion methods are compared in both subjective and objective aspects. Both qualitative and quantitative performance analysis of experimental results confirm that the proposed multi-scale decomposition image fusion method can produce high-quality HDR images.

Hydrogen sulfide detection and zebrafish imaging by a designed sensitive and selective fluorescent probe based on resorufin.

A new long-wavelength fluorescent probe 1 that could specifically identify H2S has been successfully synthesized and applied for imaging H2S in zebrafish. Probe 1 was readily prepared by featuring nitrobenzene as the recognition unit coupled to resorufin. The fluorescence off-on response is based on the fact that H2S can reduce the nitro group to an amino group, followed by the 1,6-rearrangement-elimination and the release of resorufin. By evaluating the application abilities of probe 1 in vivo and vitro, it is shown that probe 1 has high sensitivity and selectivity to H2S, low background fluorescence interference, with a low detection limit of 17.30 µM. Notably, the occurrence of the reaction can be observed by the naked eye, and the color of the solution changes from yellow to pink. More importantly, it is the first time that using paper chips as carrier to detect H2S, which lays a foundation for the practical application of detecting H2S. The excellent analysis and application capabilities of probe 1 make it an effective tool for further application in practice.

CE-MS analysis of heroin and its basic impurities using a charged polymer-protected gold nanoparticle-coated capillary.

The first application of charged polymer-protected gold nanoparticles (Au NPs) as semi-permanent capillary coating in CE-MS was presented. Poly(diallyldimethylammonium chloride) (PDDA) was the only reducing and stabilizing agent for Au NPs preparation. Stable and repeatable coating with good tolerance to 0.1 M HCl, methanol, and ACN was obtained via a simple rinsing procedure. Au NPs enhanced the coating stability toward flushing by methanol, improved the run-to-run and capillary-to-capillary repeatabilities, and improved the separation efficiency of heroin and its basic impurities for tracing geographical origins of illicit samples. Baseline resolution of eight heroin-related alkaloids was achieved on the PDDA-protected Au NPs-coated capillary under the optimum conditions: 120 mM ammonium acetate (pH 5.2) with addition of 13% methanol, separation temperature 20 degrees C, applied voltage -20 kV, and capillary effective length 60.0 cm. CE-MS analysis with run-to-run RSDs (n=5) of migration time in the range of 0.43-0.62% and RSDs (n=5) of peak area in the range of 1.49-4.68% was obtained. The established CE-MS method would offer sensitive detection and confident identification of heroin and related compounds and provide an alternative to LC-MS and GC-MS for illicit drug control.

Fragmentation pathways of heroin-related alkaloids revealed by ion trap and quadrupole time-of-flight tandem mass spectrometry.

The electrospray ionization (ESI) ion trap and quadrupole time-of-flight (QqToF) mass spectra of heroin and seven related alkaloids, i.e., morphine, codeine, O-6-monoacetylmorphine (6-MAM), thebaine, acetylcodeine, papaverine and narcotine, have been extensively investigated in this work. The ESI mass spectrometric fragmentation pathways of protonated 6-MAM, heroin, acetylcodeine, and thebaine were comprehensively elucidated for the first time with the aid of high-resolution mass spectrometry. It was found that cleavage of the piperidine ring was the featured fragmentation route of six of the compounds, although not of papaverine and narcotine. In addition, a simple high-performance liquid chromatography (HPLC)-based separation method gave baseline resolution of all eight components. This study could play an important role in the screening for these alkaloids in different matrices by HPLC coupled to tandem mass spectrometry (MS/MS).