L-homophenylalanine (L-HPA) is an important non-natural amino acid that has been used as a key intermediate for the synthesis of Puli drugs for the treatment of hypertension. At present, L-HPA is synthesized using chemical methods, which has the disadvantages of expensive raw materials, tedious steps and serious pollution. Therefore, researchers have conducted in-depth research on the enzymatic production of L-HPA. This review summarizes the research progress on the enzymatic synthesis of L-HPA, including the dehydrogenase process, the transaminase process, the hydantoinase process, and the decarboxylase process, with the hope to facilitate the industrial production of L-HPA.
Amino Acids ; Environmental Pollution ; Industry ; Protein Biosynthesis

With the development of high-throughput sequencing technology, circular RNAs (circRNAs) have gradually become a hotspot in the research on non-coding RNA. CircRNAs are produced by the covalent circularization of a downstream 3' splice donor and an upstream 5' splice acceptor through backsplicing, and they are pervasive in eukaryotic cells. CircRNAs used to be considered byproducts of false splicing, whereas an explosion of related studies in recent years has disproved this misconception. Compared with the rich studies of circRNAs in animals, the study of circRNAs in plants is still in its infancy. In this review, we introduced the discovery of plant circRNAs, the discovery of plant circRNAs, the circularization feature, expression specificity, conservation, and stability of plant circRNAs and expounded the identification tools, main types, and biogenesis mechanisms of circRNAs. Furthermore, we summarized the potential roles of plant circRNAs as microRNA (miRNA) sponges and translation templates and in response to biotic/abiotic stress, and briefed the degradation and localization of plant circRNAs. Finally, we discussed the challenges and proposed the future directions in the research on plant circRNAs.
Animals ; MicroRNAs/metabolism* ; Organelle Biogenesis ; Plants/metabolism* ; Protein Biosynthesis/physiology* ; RNA, Circular/metabolism* ; RNA, Plant/metabolism* ; Research/trends* ; Stress, Physiological/genetics*

In this research, we studied the formation of Drosophila melanogaster FADD (Fas-associated death domain-containing protein) amyloid fiber and its influence on signal transduction in IMD (Immune deficiency) signaling pathway to better understand the regulation mechanism of Drosophila innate immune signaling pathway, which will provide reference for the immune regulation in other species. First, we purified dFADD protein expressed in Escherichia coli and performed Sulfur flavin T binding and transmission electron microscopy to identify the dFADD amyloid fibers formed in vitro. Then we investigated the formation of dFADD polymers in S2 cells using SDD-AGE and confocal microscope. We also constructed dFADD mutants to find out which domain is essential to fiber formation and its effect on IMD signal transduction. Our results revealed that dFADD could be polymerized to form amyloid fiber polymers in vitro and inside the cells. Formation of fibers relies on DED (Death-effector domain) domain of dFADD, since DED domain-deleted mutant existed as a monomer. Dual luciferase reporter assay showed that intact DED domain was required for the induction of downstream antimicrobial peptides, indicating that fiber formation was the key to IMD signal transduction. Our study revealed the role of dFADD in mediating the cascade between IMD and Dredd in the IMD signaling pathway by forming amyloid fibers, suggesting an evolutionarily conserved regulatory mechanism of innate immune signaling pathway.
Animals ; Drosophila Proteins ; biosynthesis ; immunology ; Drosophila melanogaster ; immunology ; Fas-Associated Death Domain Protein ; biosynthesis ; immunology ; Immunity, Innate ; immunology ; Signal Transduction

PEGylation is considered one of the most successful techniques to improve the characteristics of protein drugs including to increase the circulating half-life of proteins in blood and to decrease their immunogenicity and antigenicity. One known PEG modification method is to attach PEG to the free amino group, typically at lysine residues or at the N-terminal amino acid with no selectivity, resulting in a heterogeneous product mixture. This lack of selectivity can present problems when a therapeutic PEGylated protein is being developed, because predictability of activity and manufacturing reproducibility are needed for regulatory approval. Enzymatic PEGylation of proteins is one route to overcome this limitation. Transglutaminases (TGase) are enzyme candidates for site-specific PEGylation. We use human interferon alpha 2a (IFN α2a) as a test case, and predict that the potential modification residues are Gln101 by computational approach as it contains 12 potential PEGylation sites. IFN α2a was PEGylated by Y shaped PEG40k-NH2 mediated by microbial transglutaminase. Our results show that the microbial transglutaminase mediated PEGylation of IFN α2a was site-specific only at the site of Gln101 in IFN α2a, yielding the single mono-conjugate PEG-Gln101-IFN α2a with a mass of 59 374.66 Da. Circular dichroism studies showed that PEG-Gln101-IFN α2a preserved the same secondary structures as native IFN α2a. As expected, the bioactivity and pharmacokinetic profile in rats of PEG-Gln101-IFN α2a revealed a significant improvement to unmodified IFN α2a, and better than PEGASYS.
Animals ; Antiviral Agents ; Humans ; Interferon alpha-2 ; metabolism ; Interferon-alpha ; biosynthesis ; pharmacokinetics ; Polyethylene Glycols ; pharmacokinetics ; Protein Structure, Secondary ; Rats ; Recombinant Proteins ; biosynthesis ; pharmacokinetics ; pharmacology ; Reproducibility of Results ; Transglutaminases ; metabolism

The imbalance of protein metabolism is the major cause of skeletal muscle atrophy, and the decrease of protein synthesis directly leads to the occurrence and development of age-related sarcopenia. The canonical role of leucyl-tRNA synthetase (LeuRS) is ligating leucine to the cognate tRNA, and thus it plays a central role in genetic coding. With the further studies of LeuRS in recent years, LeuRS has been found to control protein homeostasis in aging skeletal muscle via its non-canonical role. In this paper, we reviewed the structure and biological features of aminoacyl-tRNA synthetase and LeuRS, and summarized the recent advances in studies on the effects of LeuRS in regulating aging skeletal muscle protein synthesis as an intracellular leucine sensor. Moreover, we also analyzed the potential role of LeuRS in activation of mammalian target of rapamycin complex 1 (mTORC1) signaling transduction pathway in response to anabolic stimuli such as exercise and amino acids ingestion. This paper may provide some new ideas for the prevention, diagnosis and treatment of age-related sarcopenia.
Amino Acyl-tRNA Synthetases ; genetics ; Leucine-tRNA Ligase ; genetics ; Muscle, Skeletal ; Protein Biosynthesis

Emerging and re-emerging RNA viruses occasionally cause epidemics and pandemics worldwide, such as the on-going outbreak of the novel coronavirus SARS-CoV-2. Herein, we identified two potent inhibitors of human DHODH, S312 and S416, with favorable drug-likeness and pharmacokinetic profiles, which all showed broad-spectrum antiviral effects against various RNA viruses, including influenza A virus, Zika virus, Ebola virus, and particularly against SARS-CoV-2. Notably, S416 is reported to be the most potent inhibitor so far with an EC of 17 nmol/L and an SI value of 10,505.88 in infected cells. Our results are the first to validate that DHODH is an attractive host target through high antiviral efficacy in vivo and low virus replication in DHODH knock-out cells. This work demonstrates that both S312/S416 and old drugs (Leflunomide/Teriflunomide) with dual actions of antiviral and immuno-regulation may have clinical potentials to cure SARS-CoV-2 or other RNA viruses circulating worldwide, no matter such viruses are mutated or not.
Animals ; Antiviral Agents ; pharmacology ; therapeutic use ; Betacoronavirus ; drug effects ; physiology ; Binding Sites ; drug effects ; Cell Line ; Coronavirus Infections ; drug therapy ; virology ; Crotonates ; pharmacology ; Cytokine Release Syndrome ; drug therapy ; Drug Evaluation, Preclinical ; Gene Knockout Techniques ; Humans ; Influenza A virus ; drug effects ; Leflunomide ; pharmacology ; Mice ; Mice, Inbred BALB C ; Orthomyxoviridae Infections ; drug therapy ; Oseltamivir ; therapeutic use ; Oxidoreductases ; antagonists & inhibitors ; metabolism ; Pandemics ; Pneumonia, Viral ; drug therapy ; virology ; Protein Binding ; drug effects ; Pyrimidines ; biosynthesis ; RNA Viruses ; drug effects ; physiology ; Structure-Activity Relationship ; Toluidines ; pharmacology ; Ubiquinone ; metabolism ; Virus Replication ; drug effects

Translationally controlled tumor proteins (TCTP) and SNF1- related protein kinase (SnRK1) are conserved and widely present in eukaryotic cells. TCTP regulates cell division, plant growth and development, and mediates plant resistance against pathogen infection. SnRK1 participates in a range of physiological processes including sugar metabolism and resistance to abiotic and biotic stresses. Previous work in our laboratory demonstrated that wheat TCTP can respond to Puccinia triticina infection and induce host defense responses. In order to further investigate the mechanism of TaTCTP in wheat resistance to Puccinia triticina infection, we used TAP (tandem affinity purification) and mass spectrometry to screen the potential interactants of TaTCTP. A SNF1- related protein kinase (SnRK1) was identified as a potential interacting protein of TaTCTP. The results of yeast two-hybrid assay showed that TCTP could interact with SnRK1 in yeast, and the yeast carrying TCTP and SnRK1 could grow on SD/-Leu/-Trp/-His/-Ade (SD/-LWHA) medium. The fluorescence signal of the interaction between TCTP and SnRK1 was found to be distributed in the cytoplasm in the Bi-fluorescense complementation experiment. Co-IP experiments further showed that TCTP and SnRK1 could interact in plant cells. This study lays an important foundation for further studying the mechanism of TaTCTP in the interaction between wheat and Puccinia triticina, and it play a great influence on further improving the molecular mechanism of wheat resistant to Puccinia triticina.
Basidiomycota ; Humans ; Neoplasms ; Protein Biosynthesis ; Protein-Serine-Threonine Kinases ; Triticum

OBJECTIVE: To investigate the effects of high dose vitamin C (VC) on proliferation of breast cancer cells and to explore its mechanisms. METHODS: Human breast cancer cells Bcap37 and MDA-MB-453 were treated with VC at low dose (0.01 mmol/L), medium dose (0.10 mmol/L) and high dose (2.00 mmol/L). Cell proliferation was determined with CCK-8 assay, protein expression was evaluated by Western blot, and the secretion of lactic acid in tumor cells was detected by colorimetric method. Bcap37 cells were inoculated in nude mice, and tumor baring nude mice were intraperitoneally injected with high VC(4 g/kg, VC group, =5)or normal saline (control group, =5) for 24 d. Tumor weight and body weight were calculated. RESULTS: experiments demonstrated that high dose VC significantly inhibited cell proliferation in Bcap37 and MDA-MB-453 cells (all <0.01); the expressions of Glut1 and mTOR signaling pathway-related proteins were decreased (all <0.05); and the secretion of lactic acid was also markedly reduced (all <0.05). experiment showed that the tumor weight was decreased in mice treated with high-dose VC as compared with control group (<0.05), but no difference in body weights between two groups was observed. CONCLUSIONS High dose VC may inhibit proliferation of breast cancer cells both and through reducing glycolysis and protein synthesis.
Animals ; Ascorbic Acid ; pharmacology ; Breast Neoplasms ; drug therapy ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Glycolysis ; drug effects ; Humans ; Mice ; Mice, Nude ; Protein Biosynthesis ; drug effects

The aim of this study was to investigate the role of S100 calcium binding protein A16 (S100A16) in lipid metabolism in hepatocytes and its possible biological mechanism. HepG2 cells (human hepatoma cell line) were cultured with fatty acid to establish fatty acid culture model. The control model was cultured without fatty acid. Each model was divided into three groups and transfected with S100a16 over-expression, shRNA and vector plasmids, respectively. The concentration of triglyceride (TG) in the cells was measured by kit, and the lipid droplets was observed by oil red O staining. Immunoprecipitation and mass spectrometry were used to find the interesting proteins interacting with S100A16, and the interaction was verified by immunoprecipitation. The further mechanism was studied by Western blot and qRT-PCR. The results showed that the intracellular lipid droplet and TG concentrations in the fatty acid culture model were significantly higher than those in the control model. The accumulation of intracellular fat in the S100a16 over-expression group was significantly higher than that in the vector plasmid transfection group. There was an interaction between heat shock protein A5 (HSPA5) and S100A16. Over-expression of S100A16 up-regulated protein expression levels of HSPA5, inositol-requiring enzyme 1α (IRE1α) and pIREα1, which belong to endoplasmic reticulum stress HSPA5/IRE1α-XBP1 pathway. Meanwhile, over-expression of S100A16 up-regulated the mRNA expression levels of adipose synthesis-related gene Srebp1c, Acc and Fas. In the S100a16 shRNA plasmid transfection group, the above-mentioned protein and mRNA levels were lower than those of vector plasmid transfection group. These results suggest that S100A16 may promote lipid synthesis in HepG2 cells through endoplasmic reticulum stress HSPA5/IRE1α-XBP1 pathway.
Endoplasmic Reticulum Stress ; Endoribonucleases ; physiology ; Heat-Shock Proteins ; physiology ; Hep G2 Cells ; Humans ; Lipid Metabolism ; Protein-Serine-Threonine Kinases ; physiology ; S100 Proteins ; physiology ; Triglycerides ; biosynthesis ; X-Box Binding Protein 1 ; physiology

Noncoding RNAs (ncRNAs) have played a critical role in cellular biological functions. Recently, some peptides or proteins originating from annotated ncRNAs were identified in organism development and various diseases. Here, we briefly review several novel peptides translated by annotated ncRNAs and related key functions. In addition, we summarize the potential mechanism of bifunctional ncRNAs and propose a specific "switch" triggering the transformation from the noncoding to the coding state under certain stimuli or cellular stress. The coding properties of ncRNAs and their peptide products may provide a novel horizon in proteomic research and can be regarded as a potential therapeutic target for the treatment of various diseases.
Animals ; Calcium/metabolism* ; Humans ; Open Reading Frames ; Protein Biosynthesis ; RNA, Messenger/genetics* ; RNA, Untranslated/physiology*