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Luhong He, Jairav Xolo Desai, Jinxin Gao, Laurie B Hazeltine, Zhirui Lian, John N Calley, Christopher C Frye
Oxidation of monoclonal antibodies (mAb) is a common chemical modification with potential impact on a therapeutic protein's activity and immunogenicity. In a previous study, it was found that tryptophan oxidation (Trp-ox) levels of two mAb produced in Chinese hamster ovary (CHO) cells were significantly lowered by modifying cell culture medium/feed. In this study, transcriptome analysis by RNA-Seq was applied to further elucidate the underlying mechanism of those changes in lowering the Trp-ox levels. Cell samples from the 5L fed-batch conditions were harvested and subjected to RNA-Seq analysis...
March 15, 2018: Biotechnology Journal
Teresa Cruz-Bustos, Silvia N J Moreno, Roberto Docampo
Membrane proteins in trypanosomatids are, in general, weakly expressed and confirmation of their subcellular localization frequently requires their overexpression with epitope tags. However, overexpression can lead to mislocalization of the probes. Viswanathan et al. (Nat. Methods, 2015) described high performance tags for localization of weakly expressed proteins. We report here the use of these protein tags, named "spaghetti monster", for CRISPR /Cas9-mediated C-terminal endogenous tagging of Trypanosoma cruzi to localize two weakly expressed transient receptor potential (TRP) channels to acidic compartments...
March 15, 2018: Journal of Eukaryotic Microbiology
Ine Vandewauw, Katrien De Clercq, Marie Mulier, Katharina Held, Silvia Pinto, Nele Van Ranst, Andrei Segal, Thierry Voet, Rudi Vennekens, Katharina Zimmermann, Joris Vriens, Thomas Voets
Acute pain represents a crucial alarm signal to protect us from injury. Whereas the nociceptive neurons that convey pain signals were described more than a century ago, the molecular sensors that detect noxious thermal or mechanical insults have yet to be fully identified. Here we show that acute noxious heat sensing in mice depends on a triad of transient receptor potential (TRP) ion channels: TRPM3, TRPV1, and TRPA1. We found that robust somatosensory heat responsiveness at the cellular and behavioural levels is observed only if at least one of these TRP channels is functional...
March 14, 2018: Nature
Andrea Angeli, Fatmah A S Alasmary, Sonia Del Prete, Sameh M Osman, Zeid AlOthman, William A Donald, Clemente Capasso, Claudiu T Supuran
The activation of the δ-class carbonic anhydrase (CAs, EC from the diatom Thalassiosira weissflogii (TweCAδ) was investigated using a panel of natural and non-natural amino acids and amines. The most effective activator of TweCAδ was d-Tyr (KA of 51 nM), whereas several other amino acids and amines, such as L-His, L-Trp, d-Trp, dopamine and serotonin were submicromolar activators (KA s from 0.51 to 0.93 µM). The most ineffective activator of TweCAδ was 4-amino-l-Phe (18.9 µM), whereas d-His, l-/d-Phe, l-/d-DOPA, l-Tyr, histamine, some pyridyl-alkylamines, l-adrenaline and aminoethyl-piperazine/morpholine were moderately potent activators (KA s from 1...
December 2018: Journal of Enzyme Inhibition and Medicinal Chemistry
Juan R Martinez-Galan, Ana Verdejo, Elena Caminos
Disturbances in calcium homeostasis due to canonical transient receptor potential (TRPC) and/or store-operated calcium (SOC) channels can play a key role in a large number of brain disorders. TRPC channels are plasma membrane cation channels included in the transient receptor potential (TRP) superfamily. The most widely distributed member of the TRPC subfamily in the brain is TRPC1, which is frequently linked to group I metabotropic glutamate receptors (mGluRs) and to the components of SOC channels. Proposing TRPC/SOC channels as a therapeutic target in neurological diseases previously requires a detailed knowledge of the distribution of such molecules in the brain...
2018: Frontiers in Neuroanatomy
Wesley P Schweer, John F Patience, Eric R Burrough, Brian J Kerr, Nicholas K Gabler
Porcine reproductive and respiratory syndrome virus (PRRSV) is a significant disease in the swine industry and increasing soybean meal (SBM) consumption during this disease challenge may improve performance. Our objectives were to determine the impact of SBM level on apparent total tract (ATTD) and ileal (AID) digestibility during PRRSV infection, and to determine ileal basal endogenous losses (BEL) during PRRSV infection. Forty PRRSV negative gilts were fitted with a T-cannula in the distal ileum. Treatments were arranged in a 2 × 2 factorial with high and low SBM (HSBM, 29% vs...
March 9, 2018: Journal of Animal Science
Esam A Orabi, Ann M English
S-aromatic motifs are important noncovalent forces for protein stability and function but remain poorly understood. Hence, we performed quantum calculations at the MP2(full)/6-311++G(d,p) level on complexes between Cys (H2 S, MeSH) and Met (Me2 S) models with models of Phe (benzene, toluene), Trp (indole, 3-methylindole), Tyr (phenol, 4-methylphenol) and His (imidazole, 4-methylimidazole). The most stable gas-phase conformers exhibit binding energies of -2 to -6 kcal/mol and the S atom lies perpendicular to the ring plane...
March 13, 2018: Journal of Physical Chemistry. B
Siqi Du, Yadi Wang, Choyce A Weatherly, Kylie Holden, Daniel W Armstrong
D-amino acids are now recognized to be widely present in organisms and play essential roles in biological processes. Some D-amino acids are metabolized by D-amino acid oxidase (DAO), while D-Asp and D-Glu are metabolized by D-aspartate oxidase (DDO). In this study, levels of 22 amino acids and the enantiomeric compositions of the 19 chiral proteogenic entities have been determined in the whole brain of wild-type ddY mice (ddY/DAO+/+ ), mutant mice lacking DAO activity (ddY/DAO-/- ), and the heterozygous mice (ddY/DAO+/- ) using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS)...
March 12, 2018: Analytical and Bioanalytical Chemistry
Yang Yang, Lan Luo, Xueting Cai, Yuan Fang, Jiaqi Wang, Gang Chen, Jie Yang, Qian Zhou, Xiaoyan Sun, Xiaolan Cheng, Huaijiang Yan, Wuguang Lu, Chunping Hu, Peng Cao
Oxaliplatin-induced peripheral neuropathy (OIPN) is a severe, dose-limiting toxicity associated with cancer chemotherapy. The efficacy of antioxidant administration in OIPN is debatable, as the promising preliminary results obtained with a number of antioxidants have not been confirmed in larger clinical trials. Besides its antioxidant activity, the transcription factor, nuclear factor-erythroid 2 (NF-E2) p45-related factor 2 (Nrf2) plays a crucial role in the maintenance of mitochondrial homeostasis, and mitochondrial dysfunction is a key contributor to OIPN...
March 9, 2018: Free Radical Biology & Medicine
Xiangrong Li, Kaiwei Wang, Yanru Peng
The interaction of nanoparticles (NPs) with proteins is a topic of high relevance for the medical application of nanomaterials. In the study, a comprehensive investigation was performed for the binding properties of silver nanoparticles (AgNPs) to pepsin. The results indicate that the binding of AgNPs to pepsin may be a static quenching mechanism. Thermodynamic analysis reveals that AgNPs binds to pepsin is synergistically driven by enthalpy and entropy, and the major driving forces are hydrophobic and electrostatic interactions...
March 9, 2018: Chemico-biological Interactions
Johanna M Gostner, Eva Obermayr, Ioana E Braicu, Nicole Concin, Sven Mahner, Adriaan Vanderstichele, Jalid Sehouli, Ignace Vergote, Dietmar Fuchs, Robert Zeillinger
OBJECTIVE: Circulating tumor cells (CTCs) may represent a chronic stimulus for the immune system. In the present study we investigated the potential association of CTCs, the immune activation marker neopterin, and the ratio of kynurenine to tryptophan (Kyn/Trp) as a measure for tryptophan breakdown. METHODS: Neopterin, tryptophan and kynurenine levels were measured in plasma samples from patients with benign gynecological diseases (n=65) and with primary advanced epithelial ovarian cancer (EOC) at diagnosis (n=216) and six months after adjuvant platinum-based chemotherapy (n=45) by an enzyme-linked immunosorbent assay and high performance liquid chromatography...
March 9, 2018: Gynecologic Oncology
Zhenxing Chi, Songwen Tan, Xu Tan, Lin Zhong, Weiguo Li
The pollution of heavy metals is a severer problem for the ecosystems in waters. The toxicity of Cd2+ on phycocyanin (PC) is studied in molecular level in this work. The fluorescence quenching of PC is observed by the adding Cd2+ from 0 to 500 × 10-7  mol L-1 . From the theoretical calculation and the time-resolved fluorescence decay profiles, the fluorescence quenching of PC by Cd2+ is found to be static. The synchronous fluorescence spectra are used to study the change in amino acid residues of PC molecules, indicating that the effect of Cd2+ on the Trp of PC is more significant than the Tyr...
March 11, 2018: Environmental Science and Pollution Research International
Mickaël F El Hachmane, Charlotta S Olofsson
Ca2+ impacts a large array of cellular processes in every known cell type. In the white adipocyte, Ca2+ is involved in regulation of metabolic processes such as lipolysis, glucose uptake and hormone secretion. Although the importance of Ca2+ in control of white adipocyte function is clear, knowledge is still lacking regarding the control of dynamic Ca2+ alterations within adipocytes and mechanisms inducing intracellular Ca2+ changes remain elusive. Own work has recently demonstrated the existence of store-operated Ca2+ entry (SOCE) in lipid filled adipocytes...
March 7, 2018: Biochemical and Biophysical Research Communications
Jung Gi Lee, Jae Ha Ryu, Seon-Myung Kim, Moon-Young Park, San-Ho Kim, Young G Shin, Jong-Woo Sohn, Ha Hyung Kim, Zee-Yong Park, Jae Young Seong, Jae Il Kim
Exendin-4, a 39 amino acid peptide isolated from the saliva of the Gila monster, plays an important role in regulating glucose homeostasis, and is used clinically for the treatment of type 2 diabetes. Exendin-4 shares 53% sequence identity with the incretin hormone glucagon-like peptide 1 (GLP-1) but, unlike GLP-1, is highly resistant to proteolytic enzymes such as dipeptidyl peptidase IV (DPP-IV) and neutral endopeptidase 24.11 (NEP 24.11). Herein, we focused on the structure and function of the C-terminal Trp-cage of exendin-4, and suggest that it may be structurally required for resistance to proteolysis by NEP 24...
March 6, 2018: Biochemical Pharmacology
Alberto González, Claudio A Sáez, Bernardo Morales, Alejandra Moenne
The existence of functional Transient Receptor Potential (TRP) channels was analyzed in Ectocarpus siliculosus using agonists of human TRPs and specific antagonists of TRPA1, TRPC5, TRPM8 and TRPV; intracellular calcium was detected for 60 min. Increases in intracellular calcium were observed at 13, 29, 39 and 50-52 min, which appeared to be mediated by the activation of TRPM8/V1 at 13 min, TRPV1 at 29 min, TRPA1/V1 at 39 min and TRPA1/C5 at 50-52 min. In addition, intracellular calcium increases appear to be due to extracellular calcium entry, not requiring protein kinase activation...
March 2, 2018: Plant Physiology and Biochemistry: PPB
T Nodai, S Hitomi, K Ono, C Masaki, N Harano, A Morii, M Sago-Ito, I Ujihara, T Hibino, K Terawaki, Y Omiya, R Hosokawa, K Inenaga
Oral ulcer is the most common oral disease and leads to pain during meals and speaking, reducing the quality of life of patients. Recent evidence using animal models suggests that oral ulcers induce cyclooxygenase-dependent spontaneous pain and cyclooxygenase-independent mechanical allodynia. Endothelin-1 is upregulated in oral mucosal inflammation, although it has not been shown to induce pain in oral ulcers. In the present study, we investigated the involvement of endothelin-1 signaling with oral ulcer-induced pain using our proprietary assay system in conscious rats...
March 1, 2018: Journal of Dental Research
Won Jai Jung, Sang Yeub Lee, Sue In Choi, Byung-Keun Kim, Eun Joo Lee, Kwang Ho In, Min-Goo Lee
Airway sensory nerves are known to express several receptors and channels that are activated by exogenous and endogenous mediators that cause coughing. Toll-like receptor (TLR) s are expressed in nociceptive neurons and play an important role in neuroinflammation. However, there have been very few studies of TLR expression in lung-derived sensory neurons or their relevance to respiratory symptoms such as cough. We used the bleomycin-induced pulmonary fibrosis model to investigate the change in TLR expression in pulmonary neurons and the association of TLRs with transient receptor potential (TRP) channels in pulmonary neurons...
2018: PloS One
Justyna Maliszewska, Sonia Marcinkowska, Anna Nowakowska, Hanna Kletkiewicz, Justyna Rogalska
Some natural alkaloids, e.g. capsaicin and camphor, are known to induce a desensitization state, causing insensitivity to pain or noxious temperatures in mammals by acting on TRP receptors. Our research, for the first time, demonstrated that a phenomenon of pharmacological blockade of heat sensitivity may operate in American cockroach, Periplaneta americana (L.). We studied the escape reaction time from 50°C for American cockroaches exposed to multiple doses of different drugs affecting thermo-TRP. Capsaicin, capsazepine, and camphor induced significant changes in time spent at noxious ambient temperatures...
2018: PloS One
Lin Chen, Jing He, Salim Sazzed, Rayshawn Walker
Cryo-electron microscopy (cryo-EM) is a structure determination method for large molecular complexes. As more and more atomic structures are determined using this technique, it is becoming possible to perform statistical characterization of side-chain conformations. Two data sets were involved to characterize block lengths for each of the 18 types of amino acids. One set contains 9131 structures resolved using X-ray crystallography from density maps with better than or equal to 1.5 Å resolutions, and the other contains 237 protein structures derived from cryo-EM density maps with 2-4 Å resolutions...
March 8, 2018: Molecules: a Journal of Synthetic Chemistry and Natural Product Chemistry
Naoki Wada, Takahiro Shimizu, Nobutaka Shimizu, William C de Groat, Anthony J Kanai, Pradeep Tyagi, Hidehiro Kakizaki, Naoki Yoshimura
AIMS: To investigate the role of nerve growth factor (NGF) in lower urinary tract dysfunction in mice with spinal cord injury (SCI). METHODS: Using 4-week SCI mice, single-filling cystometry and external urethral sphincter (EUS)-electromyography were performed under an awake condition. In some SCI mice, anti-NGF antibodies (10 µg/kg/h) were administered for 1 or 2 weeks before the urodynamic study. NGF levels in the bladder and L6/S1 spinal cord were assayed by ELISA...
March 8, 2018: Neurourology and Urodynamics
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