Nitrate transformation and source tracking of Yarlung Tsangpo River using a multi-tracer approach combined with Bayesian stable isotope mixing model.

Excessive levels of nitrate nitrogen (NO3--N) could lead to ecological issues, particularly in the Yarlung Tsangpo River (YTR) region located on the Qinghai Tibet Plateau. Therefore, it is crucial to understand the fate and sources of nitrogen to facilitate pollution mitigation efforts. Herein, multiple isotopes and source resolution models were applied to analyze key transformation processes and quantify the sources of NO3-. The δ15N-NO3- and δ18O-NO3- isotopic compositions in the YTR varied between 1.23‰ and 13.64‰ and -7.88‰-11.19‰, respectively. The NO3--N concentrations varied from 0.08 to -0.86 mg/L in the dry season and 0.20-1.19 mg/L during the wet season. Nitrification remained the primary process for nitrogen transformation in both seasons. However, the wet season had a widespread effect on increasing nitrate levels, while denitrification had a limited ability to reduce nitrate. The elevated nitrate concentrations during the flood season were caused by increased release of NO3- from manure & sewage (M&S) and chemical fertilizers (CF). Future endeavors should prioritize enhancing management strategies to improve the utilization efficiency of CF and hinder the direct entry of untreated sewage into the water system.

Spatial variation of soil phosphorus in the water level fluctuation zone of the Three Gorges Reservoir: Coupling effects of elevation and artificial restoration.

The water level fluctuation zone (WLFZ) is a distinctive and important component of the reservoir ecosystem. Due to periodic inundation, the fraction, spatial distribution, and chemical reactivity of soil phosphorus (P) within the WLFZ can potentially impact the loading of P into reservoir waters. However, a detailed study of this subject is lacking. In this study, the soil P in the WLFZ of the Three Gorges Reservoir, China, was examined using a combination of chemical sequential extraction, 31P NMR, and adsorption experiments. The results of chemical sequential extraction showed that HCl-Pi constituted the largest P pool among all P forms, with a mean concentration of 338 mg/kg. The content of HCl-Pi decreased significantly toward the dam, while the content of Res-P decreased in the opposite direction. The highest contents of most P forms and total P were observed at an elevation of 160 m. 31P NMR measurements showed that NaOH-EDTA Pi detectable in WLFZ soils at 145 m, 160 m, and 175 m elevation consisted mainly of orthophosphate and pyrophosphate, while NaOH-EDTA Po contained phosphate monoesters and phosphate diesters, accounting for 1.4 % to 46.2 % of NaOH-EDTA TP. Adsorption experiments showed that soil P in the WLFZ was a potential P source for reservoir waters, with chemisorption being the dominant mechanism of P sequestration. The adsorption equilibrium concentration of WLFZ soil was lower at higher elevations (>170 m) compared to lower elevations (<150 m), exhibiting a decrease in the average maximum adsorption from 271 mg/kg to 192 mg/kg. Statistical analysis suggested that Ca and Fe content, particle size, elevation, and artificial restoration were key factors affecting the fraction and content of soil P in the WLFZ. Our findings contribute to an improved understanding of the behavior of soil P in the WLFZ of large reservoirs and its potential contribution to the reservoir waters.

Knowledge, Attitude, and Practice of Breast Cancer Patients Toward Lymphedema Complications: Cross-Sectional Study.

Breast cancer is commonly treated through surgical resection, but a common complication of the procedure is lymphedema of the upper limbs, which can significantly impact patients' daily life. This study aims to investigate the knowledge, attitude, and practice (KAP) of breast cancer patients with regard to lymphedema complications. This cross-sectional study was conducted by a self-administered questionnaire between August and October 2022 toward breast cancer patients in our Hospital of Traditional Chinese Medicine. A total of 529 breast cancer patients were enrolled, including 186 (35.16%) aged < 50 years old. Participants had moderate knowledge, attitudes, and practices with scores of 18.24 ± 3.145 (possible range: 0-30), 62.24 ± 10.260 (possible range: 17-85), and 63.27 ± 20.967 (possible range: 21-105), respectively. Multivariate logistic regression showed that high school/technical secondary school (OR = 1.880, 95% CI = 1.107-3.194, P = 0.019) and being retired (OR = 0.482, 95% CI = 0.245-0.947, P = 0.034) were independently associated with good knowledge. Knowledge (OR = 1.321, 95% CI = 1.222-1.428, P < 0.001) was independently associated with a good attitude. Furthermore, knowledge (OR = 1.262, 95% CI = 1.151-1.384, P < 0.001) and attitude (OR = 1.122, 95% CI = 1.085-1.160, P < 0.001) were independently associated with good practice. Breast cancer patients have moderate knowledge, attitudes, and practices regarding lymphedema complications. Effective education and self-management programs are needed to improve patients' KAP toward lymphedema.

Community composition and abundance of complete ammonia oxidation (comammox) bacteria in the Lancang River cascade reservoir.

The construction of the reservoir has changed the nitrogen migration and transformation processes in the river, and a large amount of sediment deposition in the reservoir may also lead to the spatial differentiation of complete ammonia oxidation (comammox) bacteria. The study investigated the abundance and diversity of comammox bacteria in the sediments of three cascade reservoirs, namely, Xiaowan, Manwan, and Nuozhadu on the Lancang River in China. In these reservoirs, the average amoA gene abundance of clade A and clade B of comammox bacteria, ammonia-oxidizing archaea (AOA), and ammonia-oxidizing bacteria (AOB) was 4.16 ± 0.85 × 105, 1.15 ± 0.33 × 105, 7.39 ± 2.31 × 104, and 3.28 ± 0.99 × 105 copies g-1, respectively. The abundance of clade A was higher than that of other ammonia oxidizing microorganisms. The spatial variation of comammox bacteria abundance differed among different reservoirs, but the spatial variation trends of the two clades of comammox bacteria in the same reservoir were similar. At each sampling point, clade A1, clade A2, and clade B coexisted, and clade A2 was usually the dominant species. The connection between comammox bacteria in the pre-dam sediments was looser than that in non-pre-dam sediments, and comammox bacteria in pre-dam sediments exhibited a simpler network structure. The main factor affecting comammox bacteria abundance was NH4+-N, while altitude, temperature, and conductivity of overlying water were the main factors affecting comammox bacteria diversity. Environmental changes caused by differences in the spatial distribution of these cascade reservoirs may be the main driver of the changes of community composition and abundance of comammox bacteria. This study confirms that the construction of cascade reservoirs results in niche spatial differentiation of comammox bacteria.

Nitrogen fixation of Cyndon dactylon: A possible strategy coping with long-term flooding in the Three Gorges Reservoir.

The Three Gorges Reservoir (TGR) is one of the largest hydropower projects around the world which greatly alters the ecological function of the original ecosystem. The riparian zone of TGR is subject to a frequent fluctuation of water level, leading to severe nitrogen loss by leaching. Cyndon dactylon, a perennial stress tolerant plant, is one of the dominant plant species in the riparian zone of TGR. The underlying mechanism that C. dactylon can survive the nitrogen limitation has been under debate. In this study, we sampled the plant tissues of C. dactylon and the surrounding soils across different water levels and seasons in the riparian zone of TGR to explore the possible strategy for them to mining nitrogen. Our study found that the C. dactylon in the TGR riparian zone had endophytic nitrogen-fixing bacteria, particularly enriched in the plant foliage. The abundance of endophytic nitrogen-fixing bacteria was significantly negatively correlated with soil ammonia, nitrate, and organic matter, and significantly positively correlated with total phosphorous and moisture content. The endophytic nitrogen-fixing bacteria in C. dactylon were highly diverse, with Proteobacteria as the main dominant genera. The mutual cooperation mode among bacterial species made the endophytic nitrogen-fixing bacteria community of C. dactylon more resilient to environmental pressure, thus more readily adapting to conditions of repeated long-term flooding in the riparian zone of the TGR.

Niche differentiation of comammox Nitrospira in sediments of the Three Gorges Reservoir typical tributaries, China.

Complete ammonia oxidizer (Comammox) can complete the whole nitrification process independently, whose niche differentiation is important guarantee for its survival and ecological function. This study investigated the niche differentiation of comammox Nitrospira in the sediments of three typical tributaries of the Three Gorges Reservoir (TGR). Clade A and clade B of comammox Nitrospira coexisted in all sampling sites simultaneously. The amoA gene abundance of clade A and B was gradually increased or decreased along the flow path of the three tributaries with obvious spatial differentiation. The amoA gene abundance of comammox Nitrospira clade A (6.36 × 103 - 5.06 × 104 copies g-1 dry sediment) was higher than that of clade B (6.26 × 102 - 6.27 × 103 copies g-1 dry sediment), and the clade A amoA gene abundance was one order of magnitude higher than that of AOA (7.24 × 102 - 6.89 × 103 copies g-1 dry sediment) and AOB (1.44 × 102 - 1.46 × 103 copies g-1 dry sediment). A significant positive correlation was observed between comammox Nitrospira clade A amoA gene abundance and flow distance (P < 0.05). The number of operational taxonomic units (OTUs) in two sub-clades of clade A accounted for the majority in different tributaries, indicating that clade A also had population differentiation among different tributaries. This study revealed that comammox Nitrospira in the sediments of TGR tributaries have niche differentiation and clade A.2 played a more crucial role in comammox Nitrospira community.

The ZFP541-KCTD19 complex is essential for pachytene progression by activating meiotic genes during mouse spermatogenesis.

Meiosis is essential for fertility in sexually reproducing species and this sophisticated process has been extensively studied. Notwithstanding these efforts, key factors involved in meiosis have not been fully characterized. In this study, we investigate the regulatory roles of zinc finger protein 541 (ZFP541) and its interacting protein potassium channel tetramerization domain containing 19 (KCTD19) in spermatogenesis. ZFP541 is expressed from leptotene to the round spermatid stage, while the expression of KCTD19 is initiated in pachytene. Depletion of Zfp541 or Kctd19 leads to infertility in male mice and delays progression from early to mid/late pachynema. In addition, Zfp541-/- spermatocytes show abnormal programmed DNA double-strand break repair, impaired crossover formation and resolution, and asynapsis of the XY chromosomes. ZFP541 interacts with KCTD19, histone deacetylase 1/2 (HDAC1/2), and deoxynucleotidyl transferase terminal-interacting protein 1 (DNTTIP1). Moreover, ZFP541 binds to and activates the expression of genes involved in meiosis and post-meiosis including Kctd19; in turn, KCTD19 promotes the transcriptional activation activity of ZFP541. Taken together, our studies reveal that the ZFP541/KCTD19 signaling complex, acting as a key transcription regulator, plays an indispensable role in male fertility by regulating pachytene progression.

Early Warning of Ischemic Stroke Based on Atherosclerosis Index Combined With Serum Markers.

CONTEXT: Ischemic stroke (IS) is a serious public health problem worldwide, threatening human life and health. Atherosclerosis is the cause of stroke. At present, there are few selective indexes that can be used to evaluate atherosclerosis in the clinic; providers rely mainly on the atherosclerotic index (AI). Disturbance of lipid metabolism is considered to be a key event leading to IS. OBJECTIVE: The purpose of this study was to discover potential biomarkers in the serum of atherosclerosis-induced IS, combined with the AI to provide early warning for the diagnosis of IS. METHODS: In this study, we used nontargeted metabolomics based on ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS) to measure the changes in serum metabolites in a group of patients with IS. To verify the reproducibility of candidate biomarkers in the population, we expanded the sample size. RESULTS: Five metabolites were identified, including sphingomyelin (18:0/14:0), 1-Methylpyrrolinium, PC (18:0/18:0), LysoPC (18:0/0:0), and PC (18: 2/18:2). The combination of these 5 metabolic markers has good diagnostic and predictive ability, and the change level of these metabolites is significantly related to IS. Our results also indicate that changes in glycerophospholipid metabolism may indicate an early risk of IS development. CONCLUSION: These findings may contribute to the development of new diagnostic methods of potential biomarkers in serum combined with the AI, thereby providing early warning for the diagnosis of atherosclerosis-induced IS, and may provide a new insights for pathogenesis in IS.

Study on Hepatotoxicity of Rhubarb Based on Metabolomics and Network Pharmacology.

BACKGROUND: Rhubarb, as a traditional Chinese medicine, is the preferred drug for the treatment of stagnation and constipation in clinical practice. It has been reported that rhubarb possesses hepatotoxicity, but its mechanism in vivo is still unclear. METHODS: In this study, the chemical components in rhubarb were identified based on UPLC-Q-TOF/MS combined with data postprocessing technology. The metabolic biomarkers obtained through metabolomics technology were related to rhubarb-induced hepatotoxicity. Furthermore, the potential targets of rhubarb-induced hepatotoxicity were obtained by network pharmacology involving the above components and metabolites. Meanwhile, GO gene enrichment analysis and KEGG pathway analysis were performed on the common targets. RESULTS: Twenty-eight components in rhubarb were identified based on UPLC-Q-TOF/MS, and 242 targets related to rhubarb ingredients were predicted. Nine metabolic biomarkers obtained through metabolomics technology were closely related to rhubarb-induced hepatotoxicity, and 282 targets of metabolites were predicted. Among them, the levels of 4 metabolites, namely dynorphin B (10-13), cervonoyl ethanolamide, lysoPE (18:2), and 3-hydroxyphenyl 2-hydroxybenzoate, significantly increased, while the levels of 5 metabolites, namely dopamine, biopterin, choline, coenzyme Q9 and P1, P4-bis (5'-uridyl) tetraphosphate significantly decreased. In addition, 166 potential targets of rhubarb-induced hepatotoxicity were obtained by network pharmacology. The KEGG pathway analysis was performed on the common targets to obtain 46 associated signaling pathways. CONCLUSION: These data suggested that rhubarb may cause liver toxicity due to its action on dopamine D1 receptor (DRD1), dopamine D2 receptor (DRD2), phosphodiesterase 4B (PDE4B), vanilloid receptor (TRPV1); transient receptor potential cation channel subfamily M member 8 (TRPM8), prostanoid EP2 receptor (PTGER2), acetylcholinesterase (ACHE), muscarinic acetylcholine receptor M3 (CHRM3) through the cAMP signaling pathway, cholinergic synapses, and inflammatory mediators to regulate TRP channels. Metabolomics technology and network pharmacology were integrated to explore rhubarb hepatotoxicity to promote the reasonable clinical application of rhubarb.

ASBT(SLC10A2): A promising target for treatment of diseases and drug discovery.

Bile acids has gradually become a new focus in various diseases, and ASBT as a transporter responsible for the reabsorption of ileal bile acids, is a key hinge associated to the bile acids-cholesterol balance and bile acids of enterohepatic circulation. The cumulative studies have also shown that ASBT is a promising target for treatment of liver, gallbladder, intestinal and metabolic diseases. This article briefly reviewed the process of bile acids enterohepatic circulation, as well as the regulations of ASBT expression, covering transcription factors, nuclear receptors and gut microbiota. In addition, the relationship between ASBT and various diseases were discussed in this paper. According to the structural classification of ASBT inhibitors, the research status of ASBT inhibitors and potential ASBT inhibitors of traditional Chinese medicine (such resveratrol, jatrorrhizine in Coptis chinensis) were summarized. This review provides a basis for the development of ASBT inhibitors and the treatment strategy of related diseases.

Techniques and Strategies for Potential Protein Target Discovery and Active Pharmaceutical Molecule Screening in a Pandemic.

Viruses remain a major challenge in the fierce fight against diseases. There have been many pandemics caused by various viruses throughout the world over the years. Recently, the global outbreak of COVID-19 has had a catastrophic impact on human health and the world economy. Antiviral drug treatment has become another essential means to overcome pandemics in addition to vaccine development. How to quickly find effective drugs that can control the development of a pandemic is a hot issue that still needs to be resolved in medical research today. To accelerate the development of drugs, it is necessary to target the key target proteins in the development of the pandemic, screen active molecules, and develop reliable methods for the identification and characterization of target proteins based on the active ingredients of drugs. This article discusses key target proteins and their biological mechanisms in the progression of COVID-19 and other major epidemics. We propose a model based on these foundations, which includes identifying potential core targets, screening potential active molecules of core targets, and verifying active molecules. This article summarizes the related innovative technologies and methods. We hope to provide a reference for the screening of drugs related to pandemics and the development of new drugs.

Asymmetric responses of spatial variation of different communities to a salinity gradient in coastal wetlands.

Various ecological communities are susceptible to the salinity gradients in coastal wetlands. Remane diagram has well described the macrozoobenthos diversity pattern along salinity gradients. Yet, further research is still needed, that is, the changes in diversity and biomass of other communities (e.g. plants, fish) along salinity gradients, and whether these changes are consistent or different among different communities. In this study, using China's Yellow River Delta wetland as a case study, we analyzed the variation of the community composition, species richness, and biomass of plant, macrozoobenthos, and fish communities along a salinity gradient from <0.5 to 30 ppt. We found that plant community composition exhibited more distinct variation along the salinity gradient than macrozoobenthos, with the least distinction for fish. Plant species richness decreased greatly along the gradient, whereas macrozoobenthos richness first decreased and then increased with increasing salinity, with the low richness occurring at a salinity of 0.9-12.3 ppt. Fish had the highest richness at a salinity of 14.8-16.0 ppt. The sum of plant, macrozoobenthos, and fish species and macrozoobenthos richness were both similar to the Remane diagram. Plants had higher biomass in low-salinity zones than in high-salinity zones, except for high biomass at a salinity of 14.8-16.0 ppt, whereas macrozoobenthos and fish showed the opposite trend. Principal-coordinate analysis showed an obvious dissimilarity map based on the composition, richness, and biomass of the plant, macrozoobenthos, and fish communities. Overall, the effects of salinity gradient differed among different communities. These findings demonstrate the asymmetric responses of different communities to salinity gradients, and have practical implications for maintaining a salinity gradient in coastal wetlands.

Establishing a rapid classification and identification method for the major triterpenoids of Alisma orientale.

INTRODUCTION: Alismatis Rhizoma (AR) has been widely used to treat various diseases. Its complex chemical composition has caused certain difficulties in the analysis of this traditional Chinese medicine. Therefore, it is necessary to establish a method for the rapid classification and identification of the chemical constituents of AR. OBJECTIVE: This article describes a method for the rapid classification and identification of major triterpenoids in AR. METHODOLOGY: The samples were analysed by ultra-performance liquid chromatography quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). The assay was performed on a Waters ACQUITY UPLC BEH C18 column (100 mm × 2.1 mm, 1.7 µm) with 0.1% formic acid in water (A), and acetonitrile (B) as mobile phase by gradient elution at a flow rate of 0.3 mL/min. In the positive ion mode, the fragment information was obtained and compared with the characteristic fragments and neutral losses described in the literature. Then, the rapid classification and identification of the chemical components from AR were achieved. RESULTS: Finally, 25 triterpene compounds of AR were identified. CONCLUSIONS: The method established in this study achieved the rapid classification and identification of chemical components in AR, which promotes the development of research methods to study the constituents of traditional Chinese medicine.

A comparison of coastal habitat restoration projects in China and the United States.

We compared coastal restoration projects in a developing country, China, and a developed country, the United States of America, both of which are facing loss and degradation of coastal habitats at similar latitudes, for the period of 1992-2014. To document the scale of coastal habitat restoration projects in the two countries, we identified 914 coastal restoration projects with an accumulated area of 300,521 acres in China, with most of our information coming from scientific papers, and 1,620 coastal restoration projects with an accumulated area of 243,064 acres in the USA, with most of our information coming from public databases. In both countries, about half the projects were in wetland habitats, but China had a greater proportion of projects in submerged habitats (43% versus 28% in the USA) and the USA a greater proportion in coastal upland habitats (21% versus 9% in China). The number of new projects steadily increased over time in China, but dropped after 2006 in the USA, although the total cost of new projects continued to increase. The number of projects in China and the total cost of projects in the USA were correlated with national GDP. Restoration projects in China used fewer techniques, had fewer partners, and took longer to complete than projects in the USA. Information about projects was incomplete, especially in China, and both countries could do more to make information publically available. We know more about project construction than project outcomes, and it is unclear whether projects are achieving their goals or whether the techniques used are optimal.

Tracing the sources of nitrate in the rivers and lakes of the southern areas of the Tibetan Plateau using dual nitrate isotopes.

Based on a quantitative analysis of nitrate concentrations, the nitrate sources and temporal variability of the rivers, lakes, and wetlands of Tibet were assessed for the first time using dual isotope technology. Water samples were collected once in July 2017 for analysis of nitrate concentration and isotopic composition. The overall values of δ15N-NO3- and δ18O-NO3- ranged from +1.8‰ to +23.0‰, and from -6.3‰ to +22.2‰ respectively. Duel isotopic composition suggested that nitrification of soil organic nitrogen was the main source of nitrate in the Yalu Tsangpo River. Furthermore, anthropogenic nitrogen inputs become more important in downstream than upstream because of intensive agricultural activities and urban input. In the rivers of the Ngari District, nitrate is mainly derived from desert deposits, manure and sewage, and chemical fertilisers. Different rivers show different characteristics of nitrate sources depending on the location, topography, landform, and climate of the river basins. Animal manure, nitrification of soil organic matter, and desert deposits are mainly responsible for the shifting of nitrate isotopic signatures in lakes, which are minimally affected by human activities. In wetlands, biological nitrification and denitrification could be the main processes of nitrogen migration and transformation. These results provide useful information in revealing the fate of nitrate in different aquatic ecosystems and different areas of Tibet.

Studies on the activation of isocitrate dehydrogenase kinase/phosphatase (AceK) by Mn2+ and Mg2.

Isocitrate dehydrogenase kinase/phosphatase (AceK) is a bifunctional enzyme with both kinase and phosphatase activities that are activated by Mg2+. We have studied the interactions of Mn2+and Mg2+ with AceK using isothermal titration calorimetry (ITC) combined with molecular docking simulations and show for the first time that Mn2+ also activates the enzyme activities. However, Mn2+ and Mg2+ exert their effects by different mechanisms. Although they have similar binding constants (of 1.11 × 105 and 0.98 × 105 M-1, respectively) for AceK and induce conformational changes of the enzyme, they do not compete for the same binding site. Instead Mn2+ appears to bind to the regulatory domain of AceK, and its effect is transmitted to the active site of the enzyme by the conformational change that it induces. The information in this study should be very useful for understanding the molecular mechanism underlying the interaction between AceK and metal ions, especially Mn2+ and Mg2+.

Analysing how plants in coastal wetlands respond to varying tidal regimes throughout their life cycles.

Important to conserve plant species in coastal wetlands throughout their life cycle. All life stages in these habitats are exposed to varying tidal cycles. It is necessary to investigate all life stages as to how they respond to varying tidal regimes. We examine three wetlands containing populations of an endangered halophyte species, each subjected to different tidal regimes: (1). wetlands completely closed to tidal cycles; (2). wetlands directly exposed to tidal cycles (3). wetlands exposed to a partially closed tidal regime. Our results showed that the most threatened stage varied between wetlands subjected to these varying tidal regimes. We hypothesis that populations of this species have adapted to these different tidal regimes. Such information is useful in developing management options for coastal wetlands and modifying future barriers restricting tidal flushing.

Loopß3αC plays an important role in the structure and function of isocitrate dehydrogenase kinase/phosphatase.

This work aims to investigate the role of the loopß3αC amino acids in the structure and function of isocitrate dehydrogenase kinase/phosphatase (AceK). The results demonstrate that the precise configuration of loopß3αC is very important for AceK structure and function: structural changes alter the affinity of the enzyme for the isocitrate dehydrogenase (ICDH), which modifies enzyme activity. Intriguingly, D340 is significant for the retention of kinase and phosphatase activities, for the conformational stability of AceK and for binding ICDH. The deletion Δ341-345 increases enzyme activity by increasing the maximum velocity and affinity for ICDH. The ß3αC loop is thus critical for the structure and function of AceK.

Simultaneous determination of five phytohormones in mungbean sprouts of China by micellar electrokinetic chromatography.

A simple and rapid micellar electrokinetic chromatography method was developed for simultaneous determination of indole-3-acetic acid, indole-3-butyric acid, gibberellic acid, abscisic acid and naphthylacetic acid in mungbean sprouts for monitoring plant growth and development. The effects of several parameters related to the separation and determination were investigated in detail. The analysis was carried out using 10 mM borax, 10 mM sodium dihydrogen phosphate, 90 mM sodium dodecyl sulfate and 5% acetonitrile as running buffer (pH 9.0). Under optimum conditions, the method demonstrated good performance concerning linearity (r, 0.9954-0.9991), precision (0.77-4.97%), the method limit of detection (LOD) and the method limit of quantitation (LOQ) (LOD, 0.011-0.177 mg/kg; LOQ, 0.035-0.590 mg/kg) and accuracy (83.62-102.56%). The results confirmed that the method is rapid, convenient and of low cost for the determination of the phytohormones.

Structural basis of calcineurin activation by calmodulin.

Calcineurin is the only known calmodulin (CaM) activated protein phosphatase, which is involved in the regulation of numerous cellular and developmental processes and in calcium-dependent signal transduction. Although commonly assumed that CaM displaces the autoinhibitory domain (AID) blocking substrate access to its active site, the structural basis underlying activation remains elusive. We have created a fused ternary complex (CBA) by covalently linking three polypeptides: CaM, calcineurin regulatory B subunit (CnB) and calcineurin catalytic A subunit (CnA). CBA catalytic activity is comparable to that of fully activated native calcineurin in the presence of CaM. The crystal structure showed virtually no structural change in the active site and no evidence of CaM despite being covalently linked. The asymmetric unit contains four molecules; two parallel CBA pairs are packed in an antiparallel mode and the large cavities in crystal packing near the calcineurin active site would easily accommodate multiple positions of AID-bound CaM. Intriguingly, the conformation of the ordered segment of AID is not altered by CaM; thus, it is the disordered part of AID, which resumes a regular α-helical conformation upon binding to CaM, which is displaced by CaM for activation. We propose that the structural basis of calcineurin activation by CaM is through displacement of the disordered fragment of AID which otherwise impedes active site access.