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Biotechnology and Applied Biochemistry

Kiyoung Kim
The clot gene is required for the biosynthesis of drosopterins, the red components of Drosophila eye pigments. However, the enzymatic role of Clot in Drosophila eye pigment formation and the molecular mechanisms underlying Clot function are not fully elucidated. In this study, we cloned and characterized Clot derived from Drosophila cDNA, and results showed that Clot exhibited ∼30% sequence identity with mammalian TRP14. In addition, we reported the three-dimensional (3D) structure of Drosophila Clot based on homology modeling...
October 28, 2017: Biotechnology and Applied Biochemistry
Chaoji Huangfu, Jinchao Zhang, Yuyuan Ma, Junting Jia, Jingxuan Li, Maomin Lv, Xiaowei Ma, Xiong Zhao, Jingang Zhang
α1-antitrypsin (AAT) is widely used to treat patients with congenital AAT deficiency. Cohn Fraction IV (Cohn F IV) is normally discarded during the manufacturing process of albumin but contains approximately 33% of plasma AAT. We established a new process for large-scale purification of AAT from it. LC-ESI-MS/MS and HPLC were applied for qualitative identification and composition analysis, respectively. Stabilizers were optimized for AAT activity protection during lyophilization and dry-heat. Virus inactivation by dry-heat and solvent/detergent (S/D) was validated on a range of viruses...
October 26, 2017: Biotechnology and Applied Biochemistry
Yunfei Gao, Xiaoqing Hu, Jianli Wang, Huazhong Li, Xiaoyuan Wang
Mycolic acid plays important role in C. glutamicum, but the key enzymes in the biosynthetic pathway of mycolic acid in C. glutamicum ATCC13869 has not been characterized. Since the locus BBD29_RS14045 in C. glutamicum ATCC13869 shows high similarity to the gene Cgl2871 which encodes Pks13, the key enzyme for synthesizing mycolic acid in C. glutamicum ATCC13032, it was deleted, resulting in the mutant WG001. Compared with the wild type ATCC1369, mycolic acid was not synthesized in WG001, but more phosphatidylglycerol and phosphatidylinositol containing longer unsaturated fatty acids were produced...
October 26, 2017: Biotechnology and Applied Biochemistry
Chih-Jung Hsu, Harsh V Jain, Abasha Williams, Julie Wang, Scott C Lute, Serge L Beaucage, Kurt A Brorson
A rapid and cost-effective transient transfection method for mammalian cells is essential for screening biopharmaceuticals in early stages of development. A library of 25 amphipathic trans-acting oligodeoxythymidine phosphorothioate triester (dTtaPS) transfection reagents, carrying positively charged and lipophilic groups, has been constructed for this purpose. High-throughput screening of the library, using an imaging cytometer and an automated microbioreactor system, has led to the identification of dTtaPS(10+) as a potent transfection reagent...
October 10, 2017: Biotechnology and Applied Biochemistry
Hani A Alhadrami
Biosensors are devices that combine a biological material with a suitable platform for detection of pathogenic organisms, carcinogenic, mutagenic and/or toxic chemicals or for reporting a biological effect. In recent years, enormous number of different types of biosensors have been constructed and developed for several medical applications. The reason for that was primarily due to the numerous advantages and applications that can be offered by biosensors. This review article has been started with demonstrating the powerful of biosensor technologies versus analytical and conventional techniques...
October 10, 2017: Biotechnology and Applied Biochemistry
Huda F AlShaibi, Farid Ahmed, Clive Buckle, Ann Cm Fowles, Jalaluddin Awlia, Marco G Cecchini, Colby L Eaton
BACKGROUND: Bone metastasis is a key event responsible for morbidity in prostate cancer patients. Interactions between prostate cancer cells and the bone microenvironment facilitate survival of tumour cells and alter bone turnover, a process that is thought to enhance the growth of metastases in this site. This study aimed to test the hypothesis that the presence of tumours cells increases TGFβ signaling in bone and that this regulates the proliferation and differentiation of osteoblastic lineage cells in metastatic sites...
October 5, 2017: Biotechnology and Applied Biochemistry
Yi Yang, Jian-Tao Yang, Xiao-Hu Chen, Ben-Gang Qin, Fu-Gui Li, Yun-Xian Chen, Li-Qiang Gu, Jia-Kai Zhu, Ping Li
We have previously demonstrated that human adipose-derived stem cells (hADSCs) can be differentiated into lymphatic endothelial like cells. The purpose of this study was to investigate the feasibility of utilizing the induced lymphatic endothelial like cells and decellularized arterial scaffold to construct the tissue-engineered lymphatic vessel. The hADSCs were isolated from adipose tissue in healthy adults and were characterized the multilineage differentiation potential. Decellularized arterial scaffold was prepared using the Triton x-100 method...
October 5, 2017: Biotechnology and Applied Biochemistry
Malgorzata Kesik-Brodacka
Since its introduction in 1982, biopharmaceutical drugs have revolutionized the treatment of a broad spectrum of diseases and are increasingly used in nearly all branches of medicine. In recent years, the biopharmaceuticals market has developed much faster than the market for all drugs and is believed to have great potential for further dynamic growth because of the tremendous demand for these drugs. Biobetters, which contain altered active pharmaceutical ingredients with enhanced efficacy, will play an important role in the development of biopharmaceuticals...
October 3, 2017: Biotechnology and Applied Biochemistry
Yuzhou Wang, Ye Li, Jianli Wang, Xiaoyuan Wang
Based on the analysis of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and mass spectrometry, flagellin FliC and a key member of the type VI secretion system Hcp1 were found to be secreted from Pseudomonas putida KT2442 cells in the late exponential phase, but in the early exponential phase only FliC was secreted. Relevant genes fleQ and rpoN were then individually deleted in P. putida KT2442, and the expression and transcription of fliC and hcp1 in these mutant strains were analyzed. In both △fleQ and △rpoN mutant cells, Hcp1 but not FliC was secreted in the late exponential phase, and it was secreted even in the early exponential phase...
October 2, 2017: Biotechnology and Applied Biochemistry
Almerinda Di Venere, Eleonora Nicolai, Federica Sinibaldi, Donato Di Pierro, Anna Maria Caccuri, Giampiero Mei
The ability of a C-terminal truncated form of TRAF2 to bind synthetic vesicles has been quantitatively studied by steady-state fluorescence energy transfer from the protein to large unilamellar vesicles (LUVs) prepared with different lipid mixtures. The dissociation constants, the free energy of binding and the average number of phospholipids interacting with truncated-TRAF2 have been evaluated from the corresponding binding curves. The results indicate that the protein strongly interacts with the lipid bilayer, preferentially in the monomeric state...
September 28, 2017: Biotechnology and Applied Biochemistry
Francesca Pintus, Annalaura Sabatucci, Mauro Maccarrone, Enrico Dainese, Rosaria Medda
This minireview focuses on a plant copper/2,4,5-trihydroxyphenyl alanine quinone amine oxidase isolated from the latex of the shrub Euphorbia characias (ELAO). This enzyme has been investigated in terms of both molecular structure and kinetic mechanism. The characterization of this enzyme allowed us to identify specific amino acids and domains that play a key role in modulating substrate access into the active site not only for ELAO but also for other plant and mammalian amine oxidases. As mammalian amine oxidases are implicated in several physiological and pathological conditions, the deep structural characterization of their active site accession mechanisms could be the starting point for the development of enzyme modulators with high therapeutic potential...
September 20, 2017: Biotechnology and Applied Biochemistry
Giovanna Di Nardo, Giuseppe Cimicata, Roberta Baravalle, Valentina Dell'Angelo, Alberto Ciaramella, Gianluca Catucci, Piero Ugliengo, Gianfranco Gilardi
Aromatase catalyzes the biosynthesis of estrogens from androgens. Owing to the physiological importance of this conversion of lipophilic substrates, the interaction with the lipid bilayer for this cytochrome P450 is crucial for its dynamics that must allow an easy access to substrates and inhibitors. Here, the aromatase-anastrozole interaction is studied by combining computational methods to identify possible access/egress routes with the protein inserted in the membrane and experimental tools aimed at the investigation of the effect of the inhibitor on the protein conformation...
September 19, 2017: Biotechnology and Applied Biochemistry
GuoQing Ying, MinJun Wang, Yu Yi, JianShu Chen, JianFeng Mei, YanLu Zhang, ShuQing Chen
An electrochemical biosensor that used an aptamer as a biological element was constructed to detect endotoxin. Biolayer interferometry was used to obtain the affinity constant of an aptamer for lipopolysaccharide, which had an equilibrium dissociation constant of 22.9 nM. The amine-terminated aptamer was then assembled on a gold electrode surface using 3-mercaptopropionic acid as an intermediate linker. The modification of the gold electrode was confirmed by cyclic voltammetry and electrochemical impedance spectroscopy...
September 9, 2017: Biotechnology and Applied Biochemistry
Qian Zhang, Li Cui, Ping Wang, Chao Deng, Qiang Wang, Xuerong Fan
Silk sericin has excellent features of antioxidant ability and good cytocompatibility; however, high water solubility and poor mechanical properties have restricted its application in biomedical fields. In this study, aimed at improving the mechanical properties of a regenerated silk sericin, the primary hydroxy groups in silk sericin were enzymatically oxidized by using laccase and 2,2',6,6'-tetramethylpiperidine-N-oxyl (TEMPO), and the generated reactive groups then reacted further with the amino groups in the sericin chains...
September 7, 2017: Biotechnology and Applied Biochemistry
Scott Huston, John Collins, Fangfang Sun, Ting Zhang, Timothy D Vaden, Y-H Percival Zhang, Jinglin Fu
A decrease in the specific activity of an enzyme is commonly observed when the enzyme is inappropriately handled or is stored over an extended period. Here, we reported a functional transition of an FMN-bound diaphorase (FMN-DI) that happened during the long-term storage process. It was found that FMN-DI did not simply lose its β-nicotinamide adenine diphosphate (NADH) dehydrogenase activity after a long-time storage, but obtained a new enzyme activity of NADH oxidase. Further mechanistic studies suggested that the alteration of the binding strength of an FMN cofactor with a DI protein could be responsible for this functional switch of the enzyme...
September 7, 2017: Biotechnology and Applied Biochemistry
Juan Manuel Ramírez-Anguita, Ismael Rodríguez-Espigares, Ramon Guixà-González, Agostino Bruno, Mariona Torrens-Fontanals, Alejandro Varela-Rial, Jana Selent
The serotonin 5-hydroxytryptamine 2A (5-HT2A ) receptor is a G-protein-coupled receptor (GPCR) relevant for the treatment of CNS disorders. In this regard, neuronal membrane composition in the brain plays a crucial role in the modulation of the receptor functioning. Since cholesterol is an essential component of neuronal membranes, we have studied its effect on the 5-HT2A receptor dynamics through all-atom MD simulations. We find that the presence of cholesterol in the membrane increases receptor conformational variability in most receptor segments...
September 6, 2017: Biotechnology and Applied Biochemistry
Laxmi Sagwan-Barkdoll, Aldwin M Anterola
CYP725A4 is a P450 enzyme from Taxus cuspidata that catalyzes the formation of taxadiene-5α-ol (T5α-ol) from taxadiene in paclitaxel biosynthesis. Past attempts expressing CYP725A4 in heterologous hosts reported the formation of 5(12)-oxa-3(11)-cyclotaxane (OCT) and/or 5(11)-oxa-3(11)-cyclotaxane (iso-OCT) instead of, or in addition to, T5α-ol. Here we report that T5α-ol is produced as a minor product by Escherichia coli expressing both taxadiene synthase (TS) and CYP725A4. The major products were OCT and iso-OCT, while trace amounts of unidentified monooxygenated taxanes were also detected by gas chromatography-mass spectrometry...
September 6, 2017: Biotechnology and Applied Biochemistry
Wei-Bin Wu, Xiao-Lei Guo, Ming-Liang Zhang, Qing-Gen Huang, Feng Qi, Jian-Zhong Huang
l-Phenylalanine is an important amino acid that is widely used in the production of food flavors and pharmaceuticals. Generally, l-phenylalanine production by engineered Escherichia coli requires a high rate of oxygen supply. However, the coexpression of Vitreoscilla hemoglobin gene (vgb), driven bya tac promoter, with the genes encoding 3-deoxy-d-arabinoheptulosonate-7-phosphate synthetase (aroF) and feedback-resistant chorismate mutase/prephenate dehydratase (pheA(fbr) ), led to increased productivity and decreased demand for aeration by E...
September 5, 2017: Biotechnology and Applied Biochemistry
Fahimeh Charbgoo, Mehrdad Behmanesh, Maryam Nikkhah
Advances in nanobiotechnology and targeting strategy could improve the delivery of therapeutic molecules into cancer cells, leading to improved treatment efficiency with minimal side effects on normal cells. To design an efficient nanocarrier, consideration of parameters that facilitate direct drug delivery into the target cells is important. We studied the effect of single wall carbon nanotubes size on their cell internalization level via the folate receptor-mediated pathway through folic acid targeting. Folate-SWNTs were covalently synthesized and characterized...
August 30, 2017: Biotechnology and Applied Biochemistry
Nadeem Ahmed, Rabbia Abbas, Mohsin Ahmad Khan, Hamid Bashir, Saad Tahir, Ahmad Usman Zafar
Interleukin-6 a pleiotropic cytokine involved in wide range biological activities. So the large scale Production of biologically active recombinant human interleukin-6 is important for its structural and functional studies. Here we report an optimized method for shake flask fermentation and simplified high yield purification procedure for the recombinant interleukin -6. This high yield expression method not only involves the optimization of the fermentation condition, but also the single step purification method as well as two-steps denaturing and one step refolding process (2DR)...
August 22, 2017: Biotechnology and Applied Biochemistry
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