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Mijanur R Molla, Alexander Böser, Akshita Rana, Karina Schwarz, Pavel A Levkin
Efficient delivery of nucleic acids into cells is of great interest in the field of cell biology and gene therapy. Despite a lot of research, transfection efficiency and structural diversity of gene-delivery vectors are still limited. A better understanding of the structure-function relationship of gene delivery vectors is also essential for the design of novel and intelligent delivery vectors, efficient in "difficult-to-transfect" cells and in vivo clinical applications. Most of the existing strategies for the synthesis of gene-delivery vectors require multiple steps and lengthy procedures...
March 20, 2018: Bioconjugate Chemistry
Seyedmehdi Hossaini Nasr, Anne Tonson, Mohammad El-Dakdouki, David C Zhu, Dalen Agnew, Robert Wiseman, Chunqi Qian, Xuefei Huang
Atherosclerosis is an inflammatory disease of arterial walls and the rupturing of atherosclerotic plaques is a major cause of heart attack and stroke. Imaging techniques that can enable the detection of atherosclerosis plaques before clinical manifestation are urgently needed. Magnetic resonance imaging (MRI) is a powerful technique to image the morphology of atherosclerotic plaques. In order to better analyze molecular processes in plaques, contrast agents that can selectively bind to plaque receptors will prove invaluable...
March 20, 2018: ACS Applied Materials & Interfaces
Lisa A Santry, Thomas M McAusland, Leonardo Susta, Geoffrey A Wood, Pierre P Major, Jim J Petrik, Byram W Bridle, Sarah K Wootton
Newcastle disease virus (NDV) is a single-stranded, negative-sense RNA virus in the Paramyxoviridae family. Although primarily an avian pathogen, NDV is a potent oncolytic virus that has been shown to be safe and effective in a variety of preclinical cancer models and human clinical trials. To produce virus for oncolytic trials, NDV is commonly amplified in embryonated chicken eggs and purified from the allantoic fluid. Conventional methods for purifying virus from allantoic fluid often result in relatively low-titer preparations containing high levels of impurities, including immunogenic chicken host cell proteins from allantoic fluid...
June 15, 2018: Molecular Therapy. Methods & Clinical Development
Arun Kumar C Huded, Pavankumar Jingade, Manoj Kumar Mishra
Isolation of high-quality RNA from coffee is challenging because of high level of polysaccharides, polyphenols and other secondary metabolites. In the present study, a rapid and efficient RNA extraction protocol from different tissues of coffee was optimized. Sufficiently high quality and quantity (225.6-454.8 µg/g) of RNA was obtained by using the optimized protocol. The presence of two distinct bands of 28S rRNA and 18S rRNA in agarose gel proved the intactness of the RNA samples. The average spectrophotometric values of the isolated RNA ranged from 1...
March 2018: 3 Biotech
Md Saddam Hossain, Takahiro Tanaka, Kazuyoshi Takagi, Junji Hayashi, Mamoru Wakayama
In the present study, we purified α-amino acid ester hydrolase (AEH) from cell-free extracts of the Stenotrophomonas maltophilia strain HS1. The approximately 70-kDa AEH from S. maltophilia HS1 (SmAEH) was homogeneous in sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) analyses, and was present as a tetramer in gel-filtration experiments. The activity of the SmAEH enzyme was then determined by monitoring the synthesis of the antihypertensive agent dipeptide isoleucyl-tryptophan (Ile-Trp) from isoleucyl methyl ester (Ile-OMe) and tryptophan (Trp)...
March 2018: 3 Biotech
Janin Lautenschläger, Eugene V Mosharov, Ellen Kanter, David Sulzer, Gabriele S Kaminski Schierle
Postnatally derived cultures of ventral mesencephalic neurons offer several crucial advantages over embryonic ventral mesencephalic cultures, including a higher content of TH-positive cells and the ability to derive cells from the substantia nigra, which contains the neurons most vulnerable to Parkinson's disease. On the other hand, these cultures are more challenging to produce consistently. Here, we provide an easy-to-implement protocol for culturing postnatal ventral mesencephalic cells from the substantia nigra (SN) and the ventral tegmental area using commercially available media, dishes, and general lab equipment, avoiding extensive material and equipment purchases...
2018: Frontiers in Cellular Neuroscience
Siddhant Bhoir, Althaf Shaik, Vijay Thiruvenkatam, Sivapriya Kirubakaran
Human Tousled-like kinases (TLKs) are highly conserved serine/threonine protein kinases responsible for cell proliferation, DNA repair, and genome surveillance. Their possible involvement in cancer via efficient DNA repair mechanisms have made them clinically relevant molecular targets for anticancer therapy. Innovative approaches in chemical biology have played a key role in validating the importance of kinases as molecular targets. However, the detailed understanding of the protein structure and the mechanisms of protein-drug interaction through biochemical and biophysical techniques demands a method for the production of an active protein of exceptional stability and purity on a large scale...
March 19, 2018: Scientific Reports
Min Zhang, Zhanqing Wang, Lili Chi, Jing Sun, Yaling Shen
Escherichia coli is an essential host for large-scale expression of heterologous polypeptides. However, further applications are limited by the formation of potential protein aggregates. In this work, we developed a novel on-column tag removal and purification system based on Fh8 hydrophobic interaction chromatography purification and ΔI-CM self-cleavage to obtain soluble tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). We evaluated several methods to improve TRAIL solubility and finally demonstrated that the Fh8 tag was a powerful solubility enhancer...
March 16, 2018: Protein Expression and Purification
Tian Zhou, Zhiyi Lin, Pavan Puligujja, Diana Palandri, James Hilaire, Mariluz Araínga, Nathan Smith, Nagsen Gautam, JoEllyn McMillan, Yazen Alnouti, Xinming Liu, Benson Edagwa, Howard E Gendelman
AIM: While the  therapeutic potential for current long-acting (LA) antiretroviral therapy (ART) is undeniable, ligand-decorated nanoformulated LA-ART could optimize drug delivery to viral reservoirs. The development of decorated ART hinges, however, on formulation processes and manufacture efficiencies. To this end, we compared manufacture and purification techniques for ligand-decorated antiretroviral drug nanocrystals. MATERIALS & METHODS: Ligand-decorated nanoparticle manufacturing was tested using folic acid (FA) nanoformulated cabotegravir...
March 19, 2018: Nanomedicine
Chiew San Fang, Kwang-Sun Kim, Dat Thinh Ha, Moon-Soo Kim, Haesik Yang
Recombinase polymerase amplification (RPA) has been combined with electrochemical detection for simple and rapid point-of-care testing. However, there are two major hindrances to this simple and rapid testing: (i) washing or purification steps are required to remove unbound labeled probes and interfering species in the sample; (ii) it is difficult to quantify double-stranded DNA (dsDNA) electrochemically by using biospecific affinity binding without dsDNA denaturation. In the present study, we describe a wash-free and rapid electrochemical method to detect RPA-amplified dsDNAs using a zinc finger protein, Zif268...
March 19, 2018: Analytical Chemistry
Lin Jin, Aaron E Engelhart, Katarzyna P Adamala, Jack W Szostak
Liposomes containing single-chain amphiphiles, particularly fatty acids, exhibit distinct properties compared to those containing diacylphospholipids due to the unique chemical properties of these amphiphiles. In particular, fatty acid liposomes enhance dynamic character, due to the relatively high solubility of single-chain amphiphiles. Similarly, liposomes containing free fatty acids are more sensitive to salt and divalent cations, due to the strong interactions between the carboxylic acid head groups and metal ions...
February 9, 2018: Journal of Visualized Experiments: JoVE
Aurélie Fossey-Jouenne, Carine Vergne-Vaxelaire, Anne Zaparucha
Amino alcohols are versatile compounds with a wide range of applications. For instance, they have been used as chiral scaffolds in organic synthesis. Their synthesis by conventional organic chemistry often requires tedious multi-step synthesis processes, with difficult control of the stereochemical outcome. We present a protocol to enzymatically synthetize amino alcohols starting from the readily available L-lysine in 48 h. This protocol combines two chemical reactions that are very difficult to conduct by conventional organic synthesis...
February 16, 2018: Journal of Visualized Experiments: JoVE
Fatima Cabral, Colton M Miller, Katrina M Kudrna, Blake E Hass, Jocelyn G Daubendiek, Brianna M Kellar, Edward N Harris
This protocol demonstrates a method for obtaining high yield and viability for mouse hepatocytes and sinusoidal endothelial cells (SECs) suitable for culturing or for obtaining cell lysates. In this protocol, the portal vein is used as the site for catheterization, rather than the vena cava, as this limits contamination of other possible cell types in the final liver preparation. No special instrumentation is required throughout the procedure. A water bath is used as a source of heat to maintain the temperature of all the buffers and solutions...
February 12, 2018: Journal of Visualized Experiments: JoVE
Meng-Die Qi, Jian Wang, Xiao-Jing Ma, Quan Zhang, Fang-Fang Wang, Ying Kang, Hui-Xin Lin, Yong Liu
Andrographolide is a main active ingredient in traditional Chinese medicine Andrographis paniculata,with a variety of pharmacological activity,widely used in clinical practice. However its biosynthetic pathway has not been resolved. Cytochrome P450 reductase provides electrons for CYP450 and plays an important role in the CYP450 catalytic process. In this study,the coding sequence of A. paniculata CPR was screened and cloned by homologous alignment,named ApCPR4. The ApCPR4 protein was obtained by prokaryotic expression...
January 2018: Zhongguo Zhong Yao za Zhi, Zhongguo Zhongyao Zazhi, China Journal of Chinese Materia Medica
Franziska Krieg, Stefan T Ochsenbein, Sergii Yakunin, Stephanie Ten Brinck, Philipp Aellen, Adrian Süess, Baptiste Clerc, Dominic Guggisberg, Olga Nazarenko, Yevhen Shynkarenko, Sudhir Kumar, Chih-Jen Shih, Ivan Infante, Maksym V Kovalenko
Colloidal lead halide perovskite nanocrystals (NCs) have recently emerged as versatile photonic sources. Their processing and optoelectronic applications are hampered by the loss of colloidal stability and structural integrity due to the facile desorption of surface capping molecules during isolation and purification. To address this issue, herein, we propose a new ligand capping strategy utilizing common and inexpensive long-chain zwitterionic molecules such as 3-( N , N -dimethyloctadecylammonio)propanesulfonate, resulting in much improved chemical durability...
March 9, 2018: ACS Energy Letters
Ge Yang, Kun Yu, Jan Kubicek, Jörg Labahn
The data presented here are related to the research article entitled "Expression, purification, and preliminary characterization of human presenilin-2" [1]. Human Presenilin-2 is the catalytic subunit of γ-secretase and a possible calcium leakage channel (Kimberly et al., 2000; Tu et al., 2006) [2], [3]. HisPS2 which was obtained by overexpression in E. coli strain C43 (DE3) was extracted by detergent solubilisation. The sample isolation efficiency by detergents and the protein identification by mass spectrometry and western blot are described...
April 2018: Data in Brief
Shweta Duggal, Noor Jailkhani, Mukul Kumar Midha, Kanury V S Rao, Ajay Kumar
The Rb protein is a tumor suppressor protein that regulates the key G1S checkpoint consequently blocking the progression of cell cycle into S-phase. Despite its pertinent role in cell cycle regulation, comprehensive information on its interacting partners across cell cycle progression is lacking. Here, we aim to submit a comprehensive set of Rb interactors as the cell progresses from G0 through G1 and S into G2 phase in HEK 293 cell line. Affinity purification of HA-tagged Rb protein along with its interactors was analyzed by mass spectrometry (AP-MS)...
April 2018: Data in Brief
Kamila Kočí, Ivana Troppová, Martin Reli, Lenka Matějová, Miroslava Edelmannová, Helena Drobná, Lada Dubnová, Anna Rokicińska, Piotr Kuśtrowski, Libor Čapek
Neodymium enriched TiO2 anatase-brookite powders were prepared by unconventional method via using pressurized hot fluids for TiO2 crystallization and purification. The photocatalysts were tested in the CH3 OH photocatalytic decomposition and they were characterized with respect to the textural (nitrogen adsorption), structural (XRD, XPS, and Raman spectroscopies), chemical (XRF), and optical (DR UV-Vis spectroscopy) and photoelectrochemical measurement. All prepared materials were nanocrystalline, had biphasic (anatase- brookite) structure and relatively large specific surface area (125 m2 ...
2018: Frontiers in Chemistry
Shuai Zhang, Yong Cao, Hao Cheng
Due to the feature of high hydrolysis, tannase is widely used in food, beverage, brewing and other fields. However, high cost in producing natural tannase makes it difficult to apply tannase to industry in a large-scale. Microbial expression systems can be used for preparing numerous amount of enzyme at low cost, so in this paper Aspergillus niger N5-5 was expressed using E. coli system. Specific primers were designed based on the Aspergillus niger N5-5 sequence N3 (GenBank, No.: KP677552), and tannase gene tan was promoted to carry 6 His tag and enzyme cutting site which contains NdeI/HindIII using PCR amplification...
December 2017: Saudi Journal of Biological Sciences
Bonnie L Russell, Samantha Gildenhuys
Bluetongue virus (BTV) is an Orbivirus that has a profound economic impact due to direct loss of livestock as well as movement bans in an attempt to prevent the spread of the disease to susceptible areas. BTV VP7, along with VP3, forms the inner capsid core of the virus where it acts as the barrier between the outer layer and the inner core housing the genetic material. Purification of BTV VP7 has proven to be problematic and expensive mainly due to its insolubility is several expression systems. To overcome this, in this paper we present a protocol for the solubilisation of BTV VP7 from inclusion bodies expressed in E...
March 15, 2018: Protein Expression and Purification
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