Protein phosphorylation database and prediction tools.

Protein phosphorylation, one of the main protein post-translational modifications, is required for regulating various life activities. Kinases and phosphatases that regulate protein phosphorylation in humans have been targeted to treat various diseases, particularly cancer. High-throughput experimental methods to discover protein phosphosites are laborious and time-consuming. The burgeoning databases and predictors provide essential infrastructure to the research community. To date, >60 publicly available phosphorylation databases and predictors each have been developed. In this review, we have comprehensively summarized the status and applicability of major online phosphorylation databases and predictors, thereby helping researchers rapidly select tools that are most suitable for their projects. Moreover, the organizational strategies and limitations of these databases and predictors have been highlighted, which may facilitate the development of better protein phosphorylation predictors in silico.

Solvent Effects Used for Optimal Simultaneous Analysis of Amino Acids via 19F NMR Spectroscopy.

19F NMR has been extensively used in simultaneous analysis of multicomponent due to its 100% natural isotope abundance, high NMR-sensitivity, and wide-range chemical shifts. The solvent effects are usually observed in NMR spectroscopy and cause large changes in 19F chemical shifts. Herein, we propose that the simultaneous analysis of a complex mixture can be achieved using solvent effects via 19F NMR spectroscopy, such as a mixture solution of amino acids (AAs). AAs are not only cell-signaling molecules, but are also considered as biomarkers of some diseases. Hence, the analysis of AAs is important for human health and the diagnosis of diseases. In this work, the key to the success of sensing 19 biogenic AAs is the use of 2-fluorobenzaldehyde (2FBA) as a highly sensitive derivatizing agent and solvent effects to produce distinguishable 19F NMR signals. As a result, the resolution of 19F NMR spectroscopy of multiple 2FBA-labeled AAs is obviously higher than other methods based on 19F NMR. Moreover, 14 and 18 AAs can be satisfactorily differentiated and unambiguously identified in different complicated media supporting the growth of mammalian cells. Furthermore, quantification of the concentration of AAs can be made, and the limit of detection reaches 10 µM. Our work provides new insights into the simultaneous analysis of a multicomponent mixture based on solvent effects by 19F NMR spectroscopy.

19F NMR enantiodiscrimination and diastereomeric purity determination of amino acids, dipeptides, and amines.

Chiral amino-group compounds are of significance for human health, such as biogenic amino acids (AAs), dipeptides, and even various drugs. Enantiospecific discrimination of these chiral compounds is vital in diagnosing diseases, identifying pathological biomarkers and enhancing pharmaceutical chemistry research. Here, we report a simple and rapid 19F NMR-based strategy to differentiate chiral AAs, dipeptides, and amines, that were derivatized with (R)-2-(2-fluorophenyl)-2-hydroxyacetic acid ((R)-2FHA). As a result, 19 proteinogenic AAs (37 isomers) as well as Gly could be concurrently resolved. Moreover, various mirror-image dipeptides, such as Ser-His, Leu-Leu, and Ala-Ala, were commendably recognized. Intriguingly, we found that the absolute configuration of AAs in the N-terminus of dipeptides decided the relative 19F chemical shifts between two enantiomers. Besides, the ability of this method for enantiodiscrimination was further demonstrated by non-AA amines, including aromatic and aliphatic amines, and even amines having chiral centers several carbons away from the amino-group. The structurally similar antibiotics, amoxicillin and ampicillin, were well discriminated. Furthermore, this method accurately determines the de or dr values of non-racemic mixtures. Therefore, our strategy provides an effective approach for 19F NMR-based enantiodiscrimination and diastereomeric purity determination of amino-group compounds.

A Practical Method for Amino Acid Analysis by LC-MS Using Precolumn Derivatization with Urea.

Amino acid (AA) analysis is important in biochemistry, food science, and clinical medicine. However, due to intrinsic limitations, AAs usually require derivatization to improve their separation and determination. Here, we present a liquid chromatography-mass spectrometry (LC-MS) method for the derivatization of AAs using the simple agent urea. The reactions proceed quantitatively under a wide range of conditions without any pretreatment steps. Urea-derivatized products (carbamoyl amino acids) of 20 AAs exhibit better separation on reversed-phase columns and increased response in a UV detector compared to underivatized ones. We applied this approach to AA analysis in complex samples using a cell culture media as a model, and it showed potential for the determination of oligopeptides. This fast, simple, and inexpensive method should be useful for AA analysis in complex samples.

trans-4-Fluoro-l-proline: A Sensitive 19F NMR Probe for the Rapid Simultaneous Enantiomeric Analysis of Multicomponent Amines.

Simultaneous enantiomeric analysis is especially important for medicine, food security, and life science. Chiral analysis of multicomponent amine mixtures still faces many challenges. Here, our work demonstrates for the first time that a novel chiral derivatizing agent CDApro based on trans-4-fluoro-l-proline (trans4Fpro) has been successfully used for the rapid simultaneous analysis of 22 chiral nonamino acid (non-AA) amines, multicomponent l/d-AAs, or mirror-image dipeptides in a mixture, as well as amines with chiral centers several carbons remote to the amino group. Furthermore, determination of enantiomeric purity and quantification of chiral amines can be made using CDApro, which serves as a robust and powerful reagent for the differentiation of multicomponent chiral amines.

Simultaneous analysis of amino acids based on discriminative 19F NMR spectroscopy.

The simultaneous analysis of amino acids (AAs) is crucial for human health, diagnosis and treatment of disease, and nutritional quality evaluation in foodstuffs. Here, we establish an easy and rapid method for the simultaneous analysis of AAs using a single reagent 2-(trifluoromethyl)benzaldehyde (oTFMBA) based on spectral-separation-enabled 19F NMR spectroscopy. oTFMBA, a highly sensitive chemosensor, is capable of analyzing 19 proteinogenic AAs or non-amino acid amines (non-AAs) in a complex mixture by adjusting the pH in a toilless way. The 19F signals of oTFMBA-labeled AAs are distributed over a wide range of ∼ 0.7 ppm, demonstrating oTFMBA with higher resolution for simultaneous analysis of AAs compared to the o-phthaldialdehyde (OPA) method (<0.6 ppm). Additionally, 12 AAs were unambiguously identified in human urine, including Asp, Ser, Gly, Thr, Glu, Arg, Ala, Val, Ile, Tyr, His, and Phe. Furthermore, our method's detection limit for AAs is 5.83 µM, illustrating sensitivity with an ∼100-fold improvement over the OPA method. This work represents an approach to the analysis of AAs or non-AAs in a complicated mixture (even biofluid) using a 19F NMR probe with high sensitivity, which is of great significance for the simultaneous analysis of multiple analytes.

α-Conotoxin Bt1.8 from Conus betulinus selectively inhibits α6/α3ß2ß3 and α3ß2 nicotinic acetylcholine receptor subtypes.

α-Conotoxins are small disulfide-rich peptides found in the venom of marine cone snails and are potent antagonists of nicotinic acetylcholine receptors (nAChRs). They are valuable pharmacological tools and have potential therapeutic applications for the treatment of chronic pain or neurological diseases and disorders. In the present study, we synthesized and functionally characterized a novel α-conotoxin Bt1.8, which was cloned from Conus betulinus. Bt1.8 selectively inhibited ACh-evoked currents in Xenopus oocytes expressing rat(r) α6/α3ß2ß3 and rα3ß2 nAChRs with an IC50 of 2.1 nM and 9.4 nM, respectively, and similar potency for human (h) α6/α3ß2ß3 and hα3ß2 nAChRs. Additionally, Bt1.8 had higher binding affinity with a slower dissociation rate for the rα6/α3ß2ß3 subtype compared to rα3ß2. The amino acid sequence of Bt1.8 is significantly different from other reported α-conotoxins targeting the two nAChR subtypes. Further Alanine scanning analyses demonstrated that residues Ile9, Leu10, Asn11, Asn12 and Asn14 are critical for its inhibitory activity at the α6/α3ß2ß3 and α3ß2 subtypes. Moreover, the NMR structure of Bt1.8 indicated the presence of a relatively larger hydrophobic zone than other α4/7-conotoxins which may explain its potent inhibition at α6/α3ß2ß3 nAChRs.

Solution structure and backbone dynamics for S1 domain of ribosomal protein S1 from Mycobacterium tuberculosis.

The pro-drug pyrazinamide is hydrolyzed to pyrazinoic acid (POA) in its use for the treatment of tuberculosis. As a molecule with bactericidal activity, POA binds to the C-terminal S1 domain of ribosomal protein S1 from Mycobacterium tuberculosis (MtRpsACTD_S1) to inhibit trans-translation. Trans-translation is a critical component of protein synthesis quality control, and is mediated by transfer-messenger RNA. Here, we have determined the solution structure of MtRpsACTD_S1(280-368), and analyzed its structural dynamics by NMR spectroscopy. The solution structure of MtRpsACTD_S1(280-368) mainly consists of five anti-parallel ß strands, two α helices, and two 310 helices. Backbone dynamics reveals that the overall structure of MtRpsACTD_S1(280-368) is rigid, but segment L326-V333 undergoes large amplitude fluctuations on picosecond to nanosecond time scales. In addition, residues V321, H322, V331 and D335 with large Rex values exhibit significant chemical or conformational exchange on microsecond to millisecond time scale. Titration of the truncated MtRpsACTD_S1(280-368) with POA shows similar characteristics to titration of MtRpsACTD_S1(280-438) with POA. In addition, diverse length fragments of MtRpsACTD_S1 show various HN resonance signals, and we find that the interaction of MtRpsA(369-481) with MtRpsACTD_S1(280-368) [Kd = (4.25 ± 0.15) mM] is responsible for the structural difference between MtRpsACTD_S1(280-368) and MtRpsACTD_S1. This work may shed light on the underlying molecular mechanism of MtRpsACTD recognizing and binding POA or mRNA, as well as the detailed mechanism of interactions between MtRpsACTD_S1(280-368) and the additional C-terminal MtRpsA(369-481).

Risk of incident chronic kidney disease in metabolically healthy obesity and metabolically unhealthy normal weight: A systematic review and meta-analysis.

Studies have reported inconsistent results about the risk of incident chronic kidney disease (CKD) in people with metabolically healthy obesity (MHO). We designed this systematic review and meta-analysis to evaluate the risk of developing CKD in people with MHO and metabolically unhealthy normal weight (MUNW). We used a predefined search strategy to retrieve eligible studies from multiple databases up to June 20, 2022. Random-effects model meta-analyses were implied to estimate the overall hazard ratio (HR) of incident CKD in obesity phenotypes. Eight prospective cohort studies, including approximately 5 million participants with a median follow-up ranging between 3 and 14 years, were included in this meta-analysis. Compared to the metabolically healthy normal weight (MHNW), the mean differences in cardiometabolic and renal risk factors in MHO, MUNW, and metabolically unhealthy obesity (MUO) were evaluated with overall HR of 1.42, 1.49, and 1.84, respectively. Compared to MHNW, the mean estimated glomerular filtration rate (eGFR) and high-density lipoprotein (HDL) were significantly lower, and low-density lipoprotein (LDL), blood pressure, blood glucose, and triglycerides were higher in MHO and MUNW. In conclusion, MHO and MUNW are not benign conditions and pose a higher risk for incident CKD. Obesity, whether in the presence or absence of metabolic health, is a risk factor for CKD.

Case report: Type 2 diabetes mellitus with plantar malignant melanoma: Report of two cases.

Malignant melanoma is a highly malignant tumor that originates from melanocytes. It has a poor prognosis and rarely occurs on the foot. Diabetic foot ulcer is one of the most serious chronic complications of diabetes. This paper reports two cases of type 2 diabetes patients with malignant melanoma on the foot. Clinicians should improve their understanding of patients with diabetes with acral malignant melanoma. When diabetic foot ulcers occur repeatedly and continue not to heal, the clinical features of the cutaneous lesions are similar to malignant melanoma, and a pathological biopsy of the lesions should be performed promptly to obtain a clear diagnosis, avoid a missed diagnosis and improve the survival rate.

Global trends in poliomyelitis research over the past 20 years: A bibliometric analysis.

Poliomyelitis is an acute infectious disease caused by poliovirus. This bibliometric analysis aims to examine the status of poliomyelitis research in the past 20 years. Information regarding polio research was obtained from the Web of Science Core Collection database. CiteSpace, VOSviewer, and Excel were used to perform visual and bibliometric analysis with respect to countries/regions, institutions, authors, journals and keywords. A total of 5,335 publications on poliomyelitis were published from 2002 to 2021. The USA was the county with the majority of publications. Additionally, the most productive institution was the Centers for Disease Control and Prevention. Sutter, RW produced the most papers and had the most co-citations. Vaccine was the journal with the most polio-related publications and citations. The most common keywords were mainly about polio immunology research ("polio," "immunization," "children," "eradication" and "vaccine"). Our study is helpful for identifying research hotspots and providing direction for future research on poliomyelitis.

A worldwide bibliometric analysis of acromegaly in the past two decades: 1999-2022.

Objectives: To conduct a bibliometric analysis to quantify and identify the current status and trends of acromegaly research in the past two decades. Materials and methods: Articles related to acromegaly that were published from 1999 to 2022 were retrieved through the Web of Science core collection (WoSCC) database. Then, they were imported into VOSviewer and CiteSpace to conduct a visualization analysis of authors, countries, institutions, citation numbers, cocitations, keywords, and references. Results: A total of 3,909 articles were identified in the study. Among them, the United States made the largest contribution to the field. Moreover, Colao A. was the most prolific author, and the University of Naples Federico II was the institution with the most publications. In addition, the Journal of Clinical Endocrinology and Metabolism was the core journal in the field. High-frequency keywords mainly included "acromegaly," "GH (Growth Hormone)," "IGH-I (Insulin-Like Growth Factor I)," "pituitary adenomas," and "octreotide." Conclusion: Studies related to acromegaly have shown stable stepwise growth over the past two decades. Interestingly, the research focus after 2016 gradually shifted from the etiology, mechanism, medications for treatment, and complications to improving prognosis and quality of life of patients with acromegaly. The current findings may provide guidance for further research in the field of acromegaly.

Global trends and current status in pheochromocytoma: a bibliometric analysis of publications in the last 20 years.

Objective: Pheochromocytoma is a rare catecholamine-producing neuroendocrine tumour originating from the chromaffin cells of the adrenal medulla or extra-adrenal paraganglia. However, there are few bibliometric studies on Pheochromocytoma. Therefore, this study was employed to summarize the global trends and current status in pheochromocytoma by bibliometric analysis. Materials and methods: The Web of Science (WOS) core collection database was searched for publications relating to pheochromocytoma from 2001 to 2021. Bibliometric analysis was used to examine the data, and Microsoft Excel was utilized to create bar graphs. In addition, VOSviewer was used to carry out co-authorship analysis, co-citation analysis and co-occurrence analysis. CiteSpace was used to analyze the keywords citation bursts. Results: A total of 8,653 publications published in 1,806 journals by 38,590 authors in 6,117 organizations from 100 countries/regions were included in our study. Among them, USA was the leading countries in terms of total publications and sum of time cited, whereas Eunice Kennedy Shriver Natl Inst Child Hlth & Hum was the leading institutions. The main publications for pheochromocytoma-related articles were Journal of clinical endocrinology &metabolism. Pacak karel and Eisenhofer Graeme were the main contributing authors. The studies on pheochromocytoma could be grouped into five clusters: Treatment, Mechanism, Etiology, Radiology and Hormones study. Moreover, the radiology study, etiology study and some specific keywords such germlines mutation, mesenchymal stem-cells, autophagy, neuroinflammation, neurotoxicity, and hemodynamic instability, may become the hot spots of future. Conclusion: Although the number of articles on pheochromocytoma has fluctuated slightly over the past 20 years, there has been an overall upward trend. In general, precision medicine research on pheochromocytoma, especially metastatic pheochromocytoma, in terms of diagnosis, treatment, and etiology will be a hot research topic in the future. This study helps to understand the research perspectives, hot spots and trends of pheochromocytoma and provide new insight and a basis for future pheochromocytoma research quickly.

Global trends and current status of amputation: Bibliometrics and visual analysis of publications from 1999 to 2021.

OBJECTIVE: To generalize the research status, hotspots, and development trends of amputation-related research. METHODS: The data from 1999 to 2021 were collected from the Web of Science core collection database, and analyzed through bibliometrics software (CiteSpace and VOSviewer) for the dual-map overlay of journals, top 25 references with the strongest citation bursts, top 25 keywords with the strongest citation bursts, and timeline of keywords. RESULTS: A total of 8,588 literature studies were involved in this study. The United States ranks the first in terms of H-index, total number of publications, and total citations. US Department of Veterans Affairs, Veterans Health Administration, and University of Washington are the major contributors to amputation. Prosthetics and Orthotics International, Archives of Physical Medicine and Rehabilitation, and Journal of Rehabilitation Research and Development are the main publication channels for articles related to amputation. Geertzen JHB, Czerniecki J, and Dijkstra PU are major contributors to amputation. In addition, research on limb salvage treatment and surgical methods for amputation will become a hotspot in the future. CONCLUSION: The total number of publications for amputation has generally increased from 1999 to 2021. Our study is beneficial for scientists to specify the research hotspot and development direction of amputation.

A novel in-situ strategy for enantiomeric discrimination and selective identification of multicomponent carboxylic acids in foods.

Chiral carboxylic acids are ubiquitous in nature and are of pivotal importance in practical applications in the field of food science and pharmaceuticals. Simultaneous enantiomeric analysis of carboxylic acids mixtures is a significant research goal. Herein, we demonstrate a new strategy involving the use of the highly selective chiral derivatizing agent (R)-2-amino-1,1,1-trifluoropropane ((R)-TFPA) for the enantiomeric discrimination and selective identification of carboxylic acids in mixtures, even structurally similar drugs. The key success of this sensing approach lies in the distinguishable 19F NMR signals of diastereoisomers formed via derivatization of the substrates by (R)-TFPA. The utility of this approach was demonstrated by simultaneously differentiating 20 chiral carboxylic acids in a complicated mixture with accurate determination of the enantiomeric excess (ee). Most importantly, our approach can be applied to food analysis, where the diverse carboxylic acids in real samples without separation were unambiguously identified, such as vinegar, yogurt, and grape.

Effect of glucagon-like peptide-1 receptor agonists on body weight in adults with obesity without diabetes mellitus-a systematic review and meta-analysis of randomized control trials.

Clinical trials have investigated the weight loss effect of glucagon-like peptide-1 receptor agonists (GLP-1 RA) in adults with obesity without diabetes mellitus, but results for weight loss efficacy were varied. We aimed to provide an up-to-date systematic review and meta-analysis for overall weight loss effect of GLP-1 RA in adults with obesity and overweight without diabetes mellitus. We retrieved eligible randomized control trials that assessed the weight loss effect of GLP-1 RA in adults (≥18 years old) without type 1/type 2 diabetes up to September 30, 2021, using Pubmed and Embase. Of 36 clinical trials assessed for eligibility, 12 trials were included, with a combined total of 11,459 participants. Compared with control groups, a more significant weight loss was seen in GLP-1 RA groups with an overall mean difference of -7.1 kg (95% CI -9.2 to -5.0) (I2  = 99%). The overall analysis results showed that GLP-1 RA improved glycemic control without increasing the risk of hypoglycemic events. Better control of blood pressure and plasma levels of LDL, HDL, and triglycerides was seen with GLP-1 RA treatment. Subgroup analysis showed greater treatment effect of semaglutide than liraglutide. Vomiting, nausea, dyspepsia, diarrhea, constipation, and abdominal pain were GLP-1 RA-associated common adverse effects.

Protein arginine phosphorylation in organisms.

Protein arginine phosphorylation (pArg), a novel molecular switch, plays a key role in regulating cellular processes. The intrinsic acid lability, hot sensitivity, and hot-alkali instability of "high-energy" phosphoamidate (PN bond) in pArg, make the investigation highly difficult and challenging. Recently, the progress in identifying prokaryotic protein arginine kinase/phosphatase and assigning hundreds of pArg proteins and phosphosites has been made, which is arousing scientists' interest and passions. It shows that pArg is tightly connected to bacteria stress response and pathogenicity, and is probably implied in human diseases. In this review, we highlight the strategies for investigation of this mysterious modification and its momentous physiological functions, and also prospect for the potentiality of drugs development targeting pArg-relative pathways.

NMR-based investigation into protein phosphorylation.

Protein phosphorylation is a major switch mechanism for cell signaling process regulation within eukaryotic cells. Abnormal phosphorylation is either a cause or consequence of human diseases. It is imperative to comprehensively delve into the relationship between phosphorylation events and cell signaling transduction. NMR spectroscopy, a potent tool for monitoring protein phosphorylation events, is applicable for identifying the phospho-sites, quantifying the kinetic rate, discovering kinase/phosphatase inhibitors and delineating phosphorylation crosstalk with other post-translational modifications. Here, we decipher the recent progress in the investigation of eukaryotic protein O-phosphorylation by NMR spectroscopy. We focus specifically on the dynamic establishment of O-phosphorylation and its role in the cell signaling processes. Simultaneously, we positively propose a strategy for the investigation of acid-labile and "high-energy" N-phosphorylation by NMR spectroscopy. We expect that the strategy could enrich the investigation of protein N-phosphorylation.

Role of metal cations and oxyanions in the regulation of protein arginine phosphatase activity of YwlE from Bacillus subtilis.

Protein arginine phosphorylation (pArg) is a relatively novel posttranslational modification. Protein arginine phosphatase YwlE negatively regulates arginine phosphorylation and consequently induces the expression of stress-response genes that are crucial for bacterial stress tolerance and pathogenic homolog Staphylococcus aureus virulence. However, little is known about the factors that affect the enzymatic activity of YwlE with the exception of the effect of oxidative stress. Herein, based on the hydrolysis of the chromogenic substrate p-nitrophenyl phosphate (pNPP) by YwlE, we investigate the role of metal cations and oxyanions in the regulation of YwlE activity. Interestingly, among the various cations that we tested, Ca2+ activates YwlE, while other cations, including Ag+, Co2+, Cd2+, and Zn2+, are inhibitory. Furthermore, as chemical analogues of phosphate, oxyanions play multiple roles in phosphatase activity. The regulatory switch Cys within the catalytic site regulates YwlE activity. Specifically, the thiol of this Cys could be alkylated by IAM (iodoacetamide) or oxidized by H2O2, resulting in enzymatic inhibition. Conversely, reducing reagents, such as DTT (dithiothreitol), ß-me (ß-mercaptoethanol), and TCEP (tris(2-carboxyethyl)phosphine) enhance YwlE activity. Additionally, as a stable analogue to pArg, pAIE binds to YwlE with a Kd of 149.1 nM and a binding stoichiometry n of 1.2 and inhibits YwlE with an IC50 of 316.3 ± 12.73 µM. The inhibition and activation of YwlE may have broad implications for the physiology, pharmacology and toxicology of metal cations and oxyanions.

A ''turn-off'' SERS assay for kinase detection based on arginine N-phosphorylation process.

A sequential "turn-off" surface enhance Raman scattering (SERS) assay platform for the detection of protein arginine kinase McsB is constructed based on arginine N-phosphorylation process. The positive charged peptide initiates the aggregation of labelled Au nanoparticles (AuNPs) to form ''Hot Spots'', resulting to a higher SERS intensity. However, the aggregation of AuNPs could also be disbanded by the addition of McsB in which the peptides are phosphorylated on arginine residues, leading to the sharp decline of SERS signal, on account of two negative charges on the phosphate group. By this strategy, a novel ''turn-off'' SERS biosensor for McsB detection based on arginine N-phosphorylation was established with high sensitivity, selectivity and simplicity. Compared with other non-enzymatic amplification methods, the sensitivity of this newly demonstrated method was improved effectively. The detection limit for McsB is 46 pM. Besides, some controlled experiments show that the assay possesses good selectivity, which is mainly decided by the specificity of kinase. Since some kinases are important biomarkers, this assay could make great contribution in biomarkers detection in complex matrices which is based on signal amplification.