Ulva (Enteromorpha) Polysaccharides and Oligosaccharides: A Potential Functional Food Source from Green-Tide-Forming Macroalgae.

The high-valued utilization of Ulva (previously known as Enteromorpha) bioresources has drawn increasing attention due to the periodic blooms of world-wide green tide. The polysaccharide is the main functional component of Ulva and exhibits various physiological activities. The Ulva oligosaccharide as the degradation product of polysaccharide not only possesses some obvious activities, but also possesses excellent solubility and bioavailability. Both Ulva polysaccharides and oligosaccharides hold promising potential in the food industry as new functional foods or food additives. Studies on Ulva polysaccharides and oligosaccharides are increasing and have been the focus of the marine bioresources field. However, the comprehensive review of this topic is still rare and do not cover the recent advances of the structure, isolation, preparation, activity and applications of Ulva polysaccharides and oligosaccharides. This review systematically summarizes and discusses the recent advances of chemical composition, extraction, purification, structure, and activity of Ulva polysaccharides as well as oligosaccharides. In addition, the potential applications as new functional food and food additives have also been considered, and these will definitely expand the applications of Ulva oligosaccharides in the food and medical fields.

Rapid Naked-Eye Tracking of On-Cell Phenotype Based on Dual-Aptamer-Weaved Cascade Assembly of Nanostructures.

Phenotypic plasticity is an emerging paradigm for providing biological and clinical insights into cancer initiation, progression, and resistance to therapy. However, it is a great challenge to track phenotypic information on live cells with high levels of sensitivity, specificity, and simplicity, when a specific cancer-cell subset is being targeted. In this work, we have successfully achieved cascade assembly of nanoparticles on the surface of specific cancer cells by designing a dual-aptamer-weaved molecular AND logic system. Taking advantage of spatial addressability, precise controllability, and targeting recognition of the nanostructure assemblies, we can precisely label the target-cell subset in a large population of similar cells and rapidly obtain phenotypic information in response to the surface changes of captured cancer cells. Without sophisticated instruments, we can know the phenotypic information on HepG2 cells in whole blood with a high level of sensitivity and rapid naked-eye tracking of on-cell phenotype changes of HepG2 cells undergoing epithelial-mesenchymal transition.

Polyvalent Biotinylated Aptamer Scaffold for Rapid and Sensitive Detection of Tau Proteins.

Biomimetic construction of artificial scaffolds has attracted increasing attention. However, the construction methods usually require redundant materials and procedures, which is inconvenient for further application. Herein, inspired by the polyvalent multifunctional structure in nature, we have designed a polyvalent biotinylated aptamer scaffold (PBAS) which can conduct analytical performance with high sensitivity and simplified procedures. To construct a PBAS, the aptamers are designed to hybridize with prepared linker probes to form polyvalent biotinylated scaffolds, which contain both multiple aptamers and signal labels. Therefore, multifunctional scaffolds can be constructed with high recognition and capture efficiency as well as significant signal amplification. Furthermore, the scaffold can be used for the assay of some disease marker proteins. By taking tau proteins as an example, the proposed aptasensor can exhibit excellent performance with a low detection limit of 153 pg mL-1 and a short assay time of 50 min, which is much better than most of the previous methods. By assays of tau proteins in both serum and artificial cerebro spinal fluid, the PBAS-based aptasensor can work well. Therefore, the scaffold may be expected to be a powerful analytical tool which may have wide applications in the detection of a variety of analytes.

Recent advances in optical aptasensor technology for amplification strategies in cancer diagnostics.

Aptamers are chemically synthetic single-stranded DNA or RNA molecules selected by molecular evolution. They have been widely used as attractive tools in biosensing and bioimaging because they can bind to a large variety of targets with high sensitivity and high affinity and specificity. As recognition elements, aptamers contribute in particular to cancer diagnostics by recognizing different cancer biomarkers, while they can also facilitate ultrasensitive detection by further employing signal amplification elements. Optical techniques have been widely used for direct and real-time monitoring of cancer-related biomolecules and bioprocesses due to the high sensitivity, quick response, and simple operation, which has greatly benefited cancer diagnostics. In this review, we highlight recent advances in optical platform-based sensing strategies for cancer diagnostics aided by aptamers. Limitations and current challenges are also discussed.

Cucurbit[8]uril-assisted peptide assembly for feasible electrochemical assay of histone acetyltransferase activity.

Accumulating findings demonstrate the importance of histone acetyltransferases (HATs) in regulating the acetylation of histones and reveal that their aberrant catalytic activities are involved in the occurrence and progress of numerous diseases. Herein, a feasible electrochemical method is proposed to assay the activity of HAT. The critical elements of the assay method are the hindrance of HAT-catalyzed acetylation against carboxypeptidase Y-catalyzed digestion and cucurbit[8]uril-assisted peptide assembly, which may recruit peptide-templated silver nanoparticles onto the electrode surface, producing significant electrochemical signals. Taking p300 as a model HAT, the assay method is validated to exhibit desirable selectivity, reproducibility, and usability in inhibitor analysis, and allow absolute activity determination in a linear range from 0.1 to 50 nM with a detection limit of 0.055 nM, which is lower than those of previous reports. Therefore, this work may provide an effective tool for HAT activity assay, which will be of great potential in HAT-related fundamental research, disease diagnosis, and drug development in the future. Graphical abstract ᅟ.

Biochemical Characterization and Degradation Pattern of a Novel Endo-Type Bifunctional Alginate Lyase AlyA from Marine Bacterium Isoptericola halotolerans.

Alginate lyases are important tools to prepare oligosaccharides with various physiological activities by degrading alginate. Particularly, the bifunctional alginate lyase can efficiently hydrolyze the polysaccharide into oligosaccharides. Herein, we cloned and identified a novel bifunctional alginate lyase, AlyA, with a high activity and broad substrate specificity from bacterium Isoptericola halotolerans NJ-05 for oligosaccharides preparation. For further applications in industry, the enzyme has been characterized and its action mode has been also elucidated. It exhibited the highest activity (7984.82 U/mg) at pH 7.5 and 55 °C. Additionally, it possessed a broad substrate specificity, showing high activities towards not only polyM (polyß-d-mannuronate) (7658.63 U/mg), but also polyG (poly α-l-guluronate) (8643.29 U/mg). Furthermore, the Km value of AlyA towards polyG (3.2 mM) was lower than that towards sodium alginate (5.6 mM) and polyM (6.7 mM). TLC (Thin Layer Chromatography) and ESI-MS (Electrospray Ionization Mass Spectrometry) were used to study the action mode of the enzyme, showing that it can hydrolyze the substrates in an endolytic manner to release a series of oligosaccharides such as disaccharide, trisaccharide, and tetrasaccharide. This study provided extended insights into the substrate recognition and degrading pattern of the alginate lyases, with a broad substrate specificity.

Enzymatic Hydrolysis of Alginate to Produce Oligosaccharides by a New Purified Endo-Type Alginate Lyase.

Enzymatic hydrolysis of sodium alginate to produce alginate oligosaccharides has drawn increasing attention due to its advantages of containing a wild reaction condition, excellent gel properties and specific products easy for purification. However, the efficient commercial enzyme tools are rarely available. A new alginate lyase with high activity (24,038 U/mg) has been purified from a newly isolated marine strain, Cellulophaga sp. NJ-1. The enzyme was most active at 50 °C and pH 8.0 and maintained stability at a broad pH range (6.0-10.0) and temperature below 40 °C. It had broad substrate specificity toward sodium alginate, heteropolymeric MG blocks (polyMG), homopolymeric M blocks (polyM) and homopolymeric G blocks (polyG), and possessed higher affinity toward polyG (15.63 mM) as well as polyMG (23.90 mM) than polyM (53.61 mM) and sodium alginate (27.21 mM). The TLC and MS spectroscopy analysis of degradation products suggested that it completely hydrolyzed sodium alginate into oligosaccharides of low degrees of polymerization (DPs). The excellent properties would make it a promising tool for full use of sodium alginate to produce oligosaccharides.

Measurement of intracellular pH changes based on DNA-templated capsid protein nanotubes.

Intracellular pH (pHi) is a fundamental modulator of cell function. Minute changes in pHi may cause great effects in many cellular activities such as metabolism and signal transduction. Herein we report an electrochemical pHi sensor based on viral-coat proteins-DNA nanotubes modified gold electrode. The sensor is pH-sensitive as a result of the pH-dependent electrochemical property of methylene blue (MB) and cell permeable owing to the polyarginine domain of the cowpea chlorotic mottle virus (CCMV) coat protein. Moreover, because the pH sensor can be translocated into cells without any further operations, the measurement of pHi changes can be greatly simplified. The pH sensor has a broad pH spectrum in the pH range from 4.0 to 9.0 and responds rapidly to the pH changes of cells, so it may hold great potential to be a valuable tool to study pH-dependent biological and pathological processes in the future.

Separation, purification and structural characterization of marine oligosaccharides: A comprehensive and systematic review of chromatographic methods.

Marine oligosaccharides have now been applied in a wide range of industry due to various kinds of physiological activities. However, the oligosaccharides with different polymeric degrees (Dps) differed in physiological activities and applicable fields. So it is promising and essential to separate, purify and structurally characterize these oligosaccharides for understanding their structure-function relationship. This review will summarize the lasted developments in the separation, purification and structural characterization of marine oligosaccharides, including the alginate oligosaccharides, carrageenan oligosaccharides, agar oligosaccharides, chitin oligosaccharides and chitosan oligosaccharides, emphasizing the successful examples of methods for separation and purification. Furthermore, an outlook for preparation of functional oligosaccharides in food biotechnology and agriculture fields is also included. This comprehensive review could definitely promote the utilization of marine functional polysaccharides for food and agriculture.

Polysaccharides and oligosaccharides originated from green algae: structure, extraction, purification, activity and applications.

With the proceeding of global warming and water eutrophication, the phenomenon of green tide has garnered significant societal interest. Consequently, researchers had increasingly focused on the potential applications of green algae biomass, particularly its polysaccharides. The polysaccharide serves as the primary active constituent of green algae and has demonstrated numerous advantageous biological activities, including antioxidant, antiviral, anticoagulant, hypolipidemic and immuno-modulatory activities. The favorable bioavailability and solubility of green algae oligosaccharides are attributed to their low molecular weight. So there has been a growing interest in researching green algae polysaccharides and oligosaccharides for the utilization of marine biological resources. This review summarized the extraction, purification, chemical structure, composition, biological activity, and potential applications prospect of polysaccharides and oligosaccharides derived from green algae. The review could be helpful for expanding the applications of polysaccharides and oligosaccharides of green algae.

Efficient degradation of the polysaccharide extracted from Enteromorpha prolifera by using a novel polysaccharide lyase family 28 enzyme with high activity.

The polysaccharides originated from Enteromorpha species exhibited versatile physiological activities and great potential in food and medicine industries. The oligosaccharides, which prepared from polysaccharide by enzymatic hydrolysis, retained the excellent activity as polysaccharide, and then revealed better solubility, bioavailability and effectiveness. However, there are few reports on Enteromorpha polysaccharide (EP)-degrading enzymes for efficient degradation of EP and high-valued utilization of Enteromorpha biomass. Herein, a novel EP-degrading enzyme, EPD1, was identified and heterologously expressed. It could efficiently hydrolyze the EP with high activity (985.755 U/mg) and exhibited optimal activity at 50 °C and a pH of 10.0. The Km value of EPD1 was 7.5512 mg·mL-1 and the Vmax value was 4.9109 µmol·min-1·mL-1. Furthermore, EPD1 demonstrated cold adaptation as evidenced by minimal activity loss following incubation at temperatures below 30 °C for 1 h. HPLC and ESI-MS analysis revealed that EPD1 could produce disaccharides, trisaccharides and tetrasaccharides as the final degradation products from EPs. In conclusion, a novel EP-degrading enzyme with high activity and excellent performance was identified and it can expand the database of EP-degrading enzymes and provide the possibility to make full use of EPs.

Gold nanoparticles and cleavage-based dual signal amplification for ultrasensitive detection of silver ions.

Silver ion (Ag(+)) is a highly toxic heavy metal ion to fungi, viruses, bacteria, and animals. Therefore, Ag(+) monitoring in water or food resources has become extraordinarily important within the scope of human health. Here, we report a gold nanoparticles and enzyme cleavage-based dual signal amplification strategy for ultrasensitive detection of Ag(+) using electrochemical techniques. This sensing platform for Ag(+) has an extremely low detection limit of 470 fM, which also has satisfactory selectivity. Thus, it can be directly used in drinking water and lake water samples. Moreover, the strategy proposed in this work may have potential to be further developed as a generalized platform for the detection of other analytes by designing new DNA sequences for specific recognition.

Ulvan and Ulva oligosaccharides: a systematic review of structure, preparation, biological activities and applications.

Ulva is one of the main green algae causing green tide disasters. Ulvan is the primarily component polysaccharide of the cell wall of Ulva and its complex structure and monosaccharide composition resulted in various biological activities. However, the high-value and effective utilization of extracted ulvan have been obstructed by limitations ranging from large molecular weight and low solubility to poor bioavailability. Ulva oligosaccharide obtained by degrading ulvan can not only ideally retain the various biological activities of ulvan very well but also effectively solve the problems of low solubility and poor bioavailability. The preparation and biological activity studies of ulvan and Ulva oligosaccharides have become a hot spot in the field of marine biological resources development research. At present, the comprehensive reviews of ulvan and Ulva oligosaccharides are still scarce. What are overviewed in this paper are the chemical composition, structure, extraction, and purification of ulvan and Ulva oligosaccharides, where research progress on the biological activities of ulvan and Ulva oligosaccharides is summarized and prospected. A theoretical and practical basis has been provided for further research on ulvan and Ulva oligosaccharides, as well as the high-value development and effective utilization of marine algae resources.

Electrochemical strategy for sensing protein phosphorylation.

We herein report a novel electrochemical method in this paper to monitor protein phosphorylation and to assay protein kinase activity based on Zr(4+) mediated signal transition and rolling circle amplification (RCA). First, substrate peptide immobilized on a gold electrode can be phosphorylated by protein kinase A. Then, Zr(4+) links phosphorylated peptide and DNA primer probe by interacting with the phosphate groups. After the introduction of the padlock probe and phi29 DNA polymerase, RCA is achieved on the surface of the electrode. As the RCA product, a very long DNA strand, may absorb a large number of electrochemical speices, [Ru(NH(3))(6)](3+), via the electrostatic interaction, localizing them onto the electrode surface, initiated by protein kinase A, a sensitive electrochemical method to assay the enzyme activity is proposed. The detection limit of the method is as low as 0.5 unit/mL, which might promise this method as a good candidate for monitoring phosphorylation in the future.

An iRGD based strategy to study electrochemically the species inside a cell.

This paper reports a method for electrical communication between the inner part of cells and an electrode with the help of iRGD peptide. Due to the enhancement of the cell penetration caused by iRGD peptide, DNA molecules, previously modified on a gold electrode surface, can be easily transfected into the cells. At the same time, doxorubicin, an anticancer drug, can also be transfected into cells with high penetration. Consequently, doxorubicin binds to DNA chains through electrostatic interaction, and the redox reaction is transferred out of the cell across the cell membrane. As a result, this work may provide a novel way to get information from inside of cells.

Evolving strategies for marine enzyme engineering: recent advances on the molecular modification of alginate lyase.

Alginate, an acidic polysaccharide, is formed by ß-d-mannuronate (M) and α-l-guluronate (G). As a type of polysaccharide lyase, alginate lyase can efficiently degrade alginate into alginate oligosaccharides, having potential applications in the food, medicine, and agriculture fields. However, the application of alginate lyase has been limited due to its low catalytic efficiency and poor temperature stability. In recent years, various structural features of alginate lyase have been determined, resulting in modification strategies that can increase the applicability of alginate lyase, making it important to summarize and discuss the current evidence. In this review, we summarized the structural features and catalytic mechanisms of alginate lyase. Molecular modification strategies, such as rational design, directed evolution, conserved domain recombination, and non-catalytic domain truncation, are also described in detail. Lastly, the application of alginate lyase is discussed. This comprehensive summary can inform future applications of alginate lyases.

Functionalization of platinum nanoparticles for electrochemical detection of nitrite.

In this work, a novel electrochemical method for nitrite detection by using functionalized platinum nanoparticles (PtNPs) is proposed. Firstly, a gold electrode is immobilized with 4-(2-aminoethyl)benzenamine. Then, PtNPs are modified with 5-[1, 2]dithiolan-3-yl-pentanoic acid [2-(naphthalene-1-ylamino)-ethyl]amide (DPAN). Consequently, in the presence of nitrite ions, Griess reaction occurs between 4-(2-aminoethyl)benzenamine on the electrode and DPAN on PtNPs, thus PtNPs are localized onto the electrode surface. So, PtNPs-electrocatalyzed reduction of H(2)O(2) can be achieved to correlate the electrochemical signal with the concentration of nitrite ions. The linear concentration range can be as wide as 10-1,000 µM, while the detection limit is as low as 5 µM. The proposed method has been also successfully applied to the detection of nitrite with the local lake water, and the result is well consistent with that obtained by UV-visible spectrophotometric method. So, this method has potential use for monitoring nitrite in drinking water supplies in the future.

Molecular Characteristics of Human Adenovirus Type 3 Circulating in Parts of China During 2014-2018.

Human adenoviruses (HAdVs) are important pathogens causing respiratory infections; 3.5-11% of childhood community-acquired pneumonia is associated with HAdV infection. Human adenovirus type 3 (HAdV-3), leading to severe morbidity and mortality, is one of the most prevalent genotype among adenoviruses responsible for acute respiratory infections (ARIs) in children in China. To identify the genetic variation of HAdV-3 in children with ARIs in China, a molecular epidemiological study was conducted. A total of 54 HAdV-3 isolated strains were obtained from children with ARIs in Beijing, Wenzhou, Shanghai, Shijiazhuang, Hangzhou, Guangzhou, and Changchun from 2014 to 2018. Thirty-two strains of which were selected for whole-genome sequencing, while the hexon, penton base, and fiber genes were sequenced for remaining strains. Bioinformatics analysis was performed on the obtained sequences. The phylogenetic analyses based on whole-genome sequences, major capsid protein genes (hexon, penton base, and fiber), and early genes (E1, E2, E3, and E4) showed that the HAdV-3 strains obtained in this study always clustered together with the reference strains from Chinese mainland, while the HAdV-3 prototype strain formed a cluster independently. Compared with the prototype strain, all strains possessed nine amino acid (AA) substitutions at neutralization antigenic epitopes of hexon. The homology models of the hexon protein of the HAdV-3 prototype and strain BJ20160214 showed that there was no evident structural change at the AA mutation sites. Two AA substitutions were found at the Arg-Gly-Asp (RGD) loop and hypervariable region 1 (HVR1) region of the penton base. A distinct AA insertion (20P) in the highly conserved PPPSY motif of the penton base that had never been reported before was observed. Recombination analysis indicated that partial regions of protein IIIa precursor, penton base, and protein VII precursor genes among all HAdV-3 strains in this study were from HAdV-7. This study showed that the genomes of the HAdV-3 strains in China were highly homologous. Some AA mutations were found at antigenic sites; however, the significance needs further study. Our data demonstrated the molecular characteristics of HAdV-3 circulating in China and was highly beneficial for further epidemiological exploration and the development of vaccines and drugs against HAdV-3.

A Multicenter Study of Viral Aetiology of Community-Acquired Pneumonia in Hospitalized Children in Chinese Mainland.

Community-acquired pneumonia (CAP) is one of the leading causes of morbidity and mortality in children worldwide. In this study, we aimed to describe the aetiology of viral infection of pediatric CAP in Chinese mainland. During November 2014 to June 2016, the prospective study was conducted in 13 hospitals. The hospitalized children under 18 years old who met the criteria for CAP were enrolled. The throat swabs or nasopharyngeal aspirates (NPAs) were collected which were then screened 18 respiratory viruses using multiplex PCR assay. Viral pathogens were present in 56.6% (1539/2721) of the enrolled cases, with the detection rate of single virus in 39.8% of the cases and multiple viruses in 16.8% of the cases. The most frequently detected virus was respiratory syncytial virus (RSV) (15.2%, 414/2721). The highest detection rate of virus was in < 6-month-age group (70.7%, 292/413). RSV, human metapneumovirus (HMPV), human parainfluenza viruses (HPIVs) and influenza B virus (Flu B) showed the similar prevalence patterns both in north and south China, but HPIVs, Flu A, human bocavirus (HBoV), human adenovirus (HAdV) and human coronaviruses (HCoVs) showed the distinct circulating patterns in north and south China. Human enterovirus/human rhinovirus (HEV/HRV) (27.6%, 27/98), HBoV (18.4%, 18/98), RSV (16.3%, 16/98) and HMPV (14.3%, 14/98) were the most commonly detected viruses in severe pneumonia cases with single virus infection. In conclusion, viral pathogens are frequently detected in pediatric CAP cases and may therefore play a vital role in the aetiology of CAP. RSV was the most important virus in hospitalized children with CAP in Chinese mainland.

Elucidation of degrading pattern and substrate recognition of a novel bifunctional alginate lyase from Flammeovirga sp. NJ-04 and its use for preparation alginate oligosaccharides.

BACKGROUND: The alginate oligosaccharides have been widely used in agriculture, medicine, and food industries due to their versatile physiological functions such as antioxidant, anticoagulant, and antineoplastic activities. The bifunctional alginate lyases can degrade the alginate polysaccharide more efficiently into alginate oligosaccharides. Therefore, it is crucial to discover new bifunctional alginate lyase for alginate oligosaccharide production. RESULTS: Herein, a novel bifunctional alginate lyase FsAlgB was cloned and identified from deep-sea bacterium Flammeovirga sp. NJ-04, which exhibited broad substrate specificity and the highest activity (1760.8 U/mg) at pH 8.0 and 40 °C. Furthermore, the K m values of FsAlgB towards polyG (0.69 mM) and polyMG (0.92 mM) were lower than that towards sodium alginate (1.28 mM) and polyM (2.06 mM). Recombinant FsAlgB was further characterized as an endolytic alginate lyase, and it can recognize the tetrasaccharide as the minimal substrate and cleave the glycosidic bonds between the subsites of - 3 and + 1. CONCLUSION: This study provided extended insights into the substrate recognition and degrading pattern of alginate lyases with broad substrate specificity.