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Protein engineering

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https://www.readbyqxmd.com/read/28647625/thermoresponsive-poly-glycidyl-ether-brushes-on-gold-surface-engineering-parameters-and-their-implication-for-cell-sheet-fabrication
#1
Silke Heinen, José Luis Cuellar-Camacho, Marie Weinhart
Thermoresponsive polymer coatings, optimized for cell adhesion and thermally-triggered cell detachment, allow the fabrication of confluent cell sheets with intact extracellular matrix. However, rational design guidelines for such coatings are rare, since temperature-triggered cell adhesion and detachment from thermoresponsive surfaces are mechanistically not well understood. Herein, we investigated the impact of molecular weight (2, 9, 24 kDa), grafting density (0.04 - 1.4 chains nm(-2)), morphology, and roughness of well-characterized thermoresponsive poly(glycidyl ether) brushes on the cell response at 37 and 20°C...
June 21, 2017: Acta Biomaterialia
https://www.readbyqxmd.com/read/28647323/mechanism-of-plant-mediated-synthesis-of-silver-nanoparticles-a-review-on-biomolecules-involved-characterisation-and-antibacterial-activity
#2
REVIEW
S Rajeshkumar, L V Bharath
Engineering a reliable and eco-accommodating methodology for the synthesis of metal nanoparticles is a crucial step in the field of nanotechnology. Plant-mediated synthesis of metal nanoparticles has been developed as a substitute to defeat the limitations of conventional synthesis approaches such as physical and chemical methods. Biomolecules, such as proteins, amino acids, enzymes, flavonoids, and terpenoids from several plant extracts have been used as a stabilising and reducing agents for the synthesis of AgNPs...
June 21, 2017: Chemico-biological Interactions
https://www.readbyqxmd.com/read/28646905/mechanistic-understanding-of-nanoparticles-interactions-with-extracellular-matrix-the-cell-and-immune-system
#3
REVIEW
Ayse Basak Engin, Dragana Nikitovic, Monica Neagu, Petra Henrich-Noack, Anca Oana Docea, Mikhail I Shtilman, Kirill Golokhvast, Aristidis M Tsatsakis
Extracellular matrix (ECM) is an extraordinarily complex and unique meshwork composed of structural proteins and glycosaminoglycans. The ECM provides essential physical scaffolding for the cellular constituents, as well as contributes to crucial biochemical signaling. Importantly, ECM is an indispensable part of all biological barriers and substantially modulates the interchange of the nanotechnology products through these barriers. The interactions of the ECM with nanoparticles (NPs) depend on the morphological characteristics of intercellular matrix and on the physical characteristics of the NPs and may be either deleterious or beneficial...
June 24, 2017: Particle and Fibre Toxicology
https://www.readbyqxmd.com/read/28646271/characterization-of-adipose-derived-stem-cells-following-burn-injury
#4
Anesh Prasai, Amina El Ayadi, Randy C Mifflin, Michael D Wetzel, Clark R Andersen, Heinz Redl, David N Herndon, Celeste C Finnerty
Severe burns induce a prolonged inflammatory response in subcutaneous adipose tissue that modulates signaling in adipose-derived stem cells (ASCs), which hold potential for healing burn wounds or generating skin substitutes. Using a 60% rat scald burn model, we conducted a series of experiments to determine which cells isolated from the adipose tissue produced inflammatory mediators and how these changes affect ASC fate and function. The stromal vascular fraction (SVF), adipocytes, and ASCs were isolated from adipose tissue at varying times up to 4 weeks postburn and from non-injured controls...
June 23, 2017: Stem Cell Reviews
https://www.readbyqxmd.com/read/28646204/optogenetic-protein-clustering-through-fluorescent-protein-tagging-and-extension-of-cry2
#5
Hyerim Park, Na Yeon Kim, Sangkyu Lee, Nury Kim, Jihoon Kim, Won Do Heo
Protein homo-oligomerization is an important molecular mechanism in many biological processes. Therefore, the ability to control protein homo-oligomerization allows the manipulation and interrogation of numerous cellular events. To achieve this, cryptochrome 2 (CRY2) from Arabidopsis thaliana has been recently utilized for blue light-dependent spatiotemporal control of protein homo-oligomerization. However, limited knowledge on molecular characteristics of CRY2 obscures its widespread applications. Here, we identify important determinants for efficient cryptochrome 2 clustering and introduce a new CRY2 module, named ''CRY2clust'', to induce rapid and efficient homo-oligomerization of target proteins by employing diverse fluorescent proteins and an extremely short peptide...
June 23, 2017: Nature Communications
https://www.readbyqxmd.com/read/28645809/enhanced-in-vivo-retention-of-low-dose-bmp-2-via-heparin-microparticle-delivery-does-not-accelerate-bone-healing-in-a-critically-sized-femoral-defect
#6
Marian H Hettiaratchi, Tel Rouse, Catherine Chou, Laxminarayanan Krishnan, Hazel Y Stevens, Mon-Tzu A Li, Todd C McDevitt, Robert E Guldberg
Bone morphogenetic protein-2 (BMP-2) is an osteoinductive growth factor used clinically to induce bone regeneration and fusion. Some complications associated with BMP-2 treatment have been attributed to rapid release of BMP-2 from conventional collagen scaffolds, motivating the development of tunable sustained-release strategies. We incorporated BMP-2-binding heparin microparticles (HMPs) into a hydrogel scaffold to improve spatiotemporal control of BMP-2 delivery to large bone defects. HMPs pre-loaded with BMP-2 were mixed into alginate hydrogels and compared to hydrogels containing BMP-2 alone...
June 20, 2017: Acta Biomaterialia
https://www.readbyqxmd.com/read/28645099/harnessing-the-natural-diversity-and-in-vitro-evolution-of-cas9-to-expand-the-genome-editing-toolbox
#7
REVIEW
Tautvydas Karvelis, Giedrius Gasiunas, Virginijus Siksnys
In the past few years, the Cas9 endonuclease from the type II CRISPR-Cas bacterial antiviral defense system has revolutionized the genome editing field. Guided by an RNA molecule, Cas9 can be reprogrammed to target almost any DNA sequence: the only limitation being the short nucleotide sequence in the vicinity of the target, termed the PAM, which is characteristic for each Cas9 protein. Streptococcus pyogenes Cas9 which recognizes the NGG PAM is currently most widely used for genome manipulation. However, Cas9 orthologues and engineered Cas9 variants offer expanded genome targeting capabilities, improved specificity and biochemical properties...
June 20, 2017: Current Opinion in Microbiology
https://www.readbyqxmd.com/read/28644828/a-mutant-of-chlamydomonas-without-lhcsr-maintains-high-rates-of-photosynthesis-but-has-reduced-cell-division-rates-in-sinusoidal-light-conditions
#8
Michael Cantrell, Graham Peers
The LHCSR protein belongs to the light harvesting complex family of pigment-binding proteins found in oxygenic photoautotrophs. Previous studies have shown that this complex is required for the rapid induction and relaxation of excess light energy dissipation in a wide range of eukaryotic algae and moss. The ability of cells to rapidly regulate light harvesting between this dissipation state and one favoring photochemistry is believed to be important for reducing oxidative stress and maintaining high photosynthetic efficiency in a rapidly changing light environment...
2017: PloS One
https://www.readbyqxmd.com/read/28644714/genome-mining-in-silico-validation-and-phase-selection-of-a-novel-aldo-keto-reductase-from-candida-glabratea-for-biotransformation
#9
Souvik Basak, Nanda Gopal Sahoo, Angayar K Pavanasam
Previously, we published cloning, over-expression, characterization and subsequent exploitation of a carbonyl reductase (cr) gene, belonging to general family aldo-keto reductase from Candida glabrata CBS138 to convert keto ester (COBE) to a chiral alcohol (ethyl-4-chloro-3-hydroxybutanoate or CHBE). Exploiting global transcription factor CRP, rDNA and transporter engineering, we have improved batch production of CHBE by trinomial bioengineering. Herein, we present the exploration of cr gene in Candida glabrata CBS138 through genome miningapproach, in silico validation of its activity and selection of its biocatalytic phase...
June 23, 2017: Bioengineered
https://www.readbyqxmd.com/read/28644480/a-polymer-nanoparticle-with-engineered-affinity-for-a-vascular-endothelial-growth-factor-vegf165
#10
Hiroyuki Koide, Keiichi Yoshimatsu, Yu Hoshino, Shih-Hui Lee, Ai Okajima, Saki Ariizumi, Yudai Narita, Yusuke Yonamine, Adam C Weisman, Yuri Nishimura, Naoto Oku, Yoshiko Miura, Kenneth J Shea
Protein affinity reagents are widely used in basic research, diagnostics and separations and for clinical applications, the most common of which are antibodies. However, they often suffer from high cost, and difficulties in their development, production and storage. Here we show that a synthetic polymer nanoparticle (NP) can be engineered to have many of the functions of a protein affinity reagent. Polymer NPs with nM affinity to a key vascular endothelial growth factor (VEGF165) inhibit binding of the signalling protein to its receptor VEGFR-2, preventing receptor phosphorylation and downstream VEGF165-dependent endothelial cell migration and invasion into the extracellular matrix...
July 2017: Nature Chemistry
https://www.readbyqxmd.com/read/28644476/enantioselective-intermolecular-benzylic-c-h-amination-catalysed-by-an-engineered-iron-haem-enzyme
#11
Christopher K Prier, Ruijie K Zhang, Andrew R Buller, Sabine Brinkmann-Chen, Frances H Arnold
C-H bonds are ubiquitous structural units of organic molecules. Although these bonds are generally considered to be chemically inert, the recent emergence of methods for C-H functionalization promises to transform the way synthetic chemistry is performed. The intermolecular amination of C-H bonds represents a particularly desirable and challenging transformation for which no efficient, highly selective, and renewable catalysts exist. Here we report the directed evolution of an iron-containing enzymatic catalyst-based on a cytochrome P450 monooxygenase-for the highly enantioselective intermolecular amination of benzylic C-H bonds...
July 2017: Nature Chemistry
https://www.readbyqxmd.com/read/28643924/exosomes-novel-regulators-of-bone-remodelling-and-potential-therapeutic-agents-for-orthodontics
#12
REVIEW
L S Holliday, K P McHugh, J Zuo, J I Aguirre, J K Neubert, W J Rody
Recent studies suggest that exosomes are involved in intercellular communication required for the maintenance of healthy bone. Exosomes are small (30-150 nm in diameter) extracellular vesicles that are formed in multivesicular bodies and are released from cells as the multivesicular bodies fuse with the plasma membrane. Regulatory exosomes have the capacity to exert profound control over target cells. They can stimulate plasma membrane receptors and are also internalized by the target cell delivering proteins, lipids, small molecules and functional RNAs from the cell of origin...
June 2017: Orthodontics & Craniofacial Research
https://www.readbyqxmd.com/read/28643483/cell-engineered-nanovesicle-as-a-surrogate-inducer-of-contact-dependent-stimuli
#13
Junho Kim, Chugmin Han, Wonju Jo, Sehong Kang, Siwoo Cho, Dayeong Jeong, Yong Song Gho, Jaesung Park
Heterotypic interactions between cells are crucial in various biological phenomena. Particularly, stimuli that regulate embryonic stem cell (ESC) fate are often provided from neighboring cells. However, except for feeder cultures, no practical methods are identified that can provide ESCs with contact-dependent cell stimuli. To induce contact-dependent cell stimuli in the absence of living cells, a novel method that utilizes cell-engineered nanovesicles (CNVs) that are made by extruding living cells through microporous membranes is described...
June 23, 2017: Advanced Healthcare Materials
https://www.readbyqxmd.com/read/28643418/protein-engineering-of-recombinant-human-bone-morphogenetic-protein-2-with-higher-interaction-with-ca-phosphate-based-scaffold-usedfor-osteogenesis
#14
Mohammad Bayat, Sahar Shojaei, Naghmeh Bahrami, Abdol Reza Mohamadnia, Pegah Shojaei, Naeim Bahrami
The aim of the present studywas to assess the recombinant bonemorphogenetic protein 2 (RHBMP-2)with higher substantively and solubility for use in calcium phosphate scaffolds for better release in differentiation of mesenchymal stem cells to osteoblast cells. By using bioinformatics tools,2 mutations (p. L10D and p. S12E) werechosen and applied in BMP2 CDS sequenceto increase interaction with calcium derived composite. The new recombinant mutated sequence (BMP2(mut) ) was synthesized and then subcloned to expression vector pBV220...
June 23, 2017: Journal of Biomedical Materials Research. Part A
https://www.readbyqxmd.com/read/28643382/synthesis-and-biological-evaluation-of-arginyl-diosgenin-conjugate-as-a-potential-bone-tissue-engineering-agent
#15
Ai-Mei Liao, Hana Jung, Ji Won Yu, Dong Hee Lee, Sang-Soo Park, Bangrong Cai, ChangJu Chun
Water-soluble arginyl-diosgenin (Arg-DG) conjugate was designed, synthesized, and evaluated for a biological activity. The Arg-DG conjugate was characterized using FT-IR, (1) H NMR, (13) C NMR, and HPLC-MS analyses, followed by a biological activity evaluation. Compared with DG, the Arg-DG conjugate showed a decreased cytotoxicity against L929 cells and an increased anti-proliferative activity against hepatocellular cells. The safety of the Arg-DG conjugate was confirmed using the highly sensitive Alamar Blue assay, which indicated that it increased the cellular metabolic activity at suitable concentrations...
June 23, 2017: Chemical Biology & Drug Design
https://www.readbyqxmd.com/read/28643343/alpha-beta-hydrolases-a-unique-structural-motif-coordinates-catalytic-acid-residue-in-40-protein-fold-families
#16
Polytimi S Dimitriou, Alexander Denesyuk, Seiji Takahashi, Satoshi Yamashita, Mark S Johnson, Toru Nakayama, Konstantin Denessiouk
The alpha/beta-hydrolases (ABH) are a family of acid-base-nucleophile catalytic triad enzymes with a common fold, but using a wide variety of substrates, having different pH optima, catalyzing unique catalytic reactions and often showing improved chemical and thermo stability. The ABH enzymes are prime targets for protein engineering. Here, we have classified active sites from 51 representative members of 40 structural ABH fold families into eight distinct conserved geometries. We demonstrate the occurrence of a common structural motif, the catalytic acid zone, at the catalytic triad acid turn...
June 23, 2017: Proteins
https://www.readbyqxmd.com/read/28643248/computational-prediction-of-crispr-cas9-target-sites-reveals-potential-off-target-risks-in-human-and-mouse
#17
Qingbo Wang, Kumiko Ui-Tei
The clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated (Cas) system is a prominent genome engineering technology. In the CRISPR/Cas system, the RNA-guided endonuclease Cas protein introduces a DNA double-stranded break at the genome position recognized by a guide RNA (gRNA) based on complementary base-pairing of about 20-nucleotides in length. The 8- or 12-mer gRNA sequence in the proximal region is especially important for target recognition, and the genes with sequence complementarity to such regions are often disrupted...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28643153/a-3e8-scfv-cys-ir800-conjugate-targeting-tag-72-in-an-orthotopic-colorectal-cancer-model
#18
Li Gong, Haiming Ding, Nicholas E Long, Brandon J Sullivan, Edward W Martin, Thomas J Magliery, Michael F Tweedle
PURPOSE: Optical surgical navigation (OSN) will be a potent tool to help surgeons more accurately and efficiently remove tumors. The purpose of this study was to evaluate a novel humanized 3E8 antibody (3E8 MAb) fragment site-specifically conjugated with IR800, 3E8.scFv.Cys-IR800, as a potential OSN agent to target colorectal adenocarcinoma. PROCEDURES: An engineered single-chain variable fragment of 3E8 MAb (targeted to TAG-72), appending a C-terminal cysteine residue (3E8...
June 22, 2017: Molecular Imaging and Biology: MIB: the Official Publication of the Academy of Molecular Imaging
https://www.readbyqxmd.com/read/28642611/engineered-newcastle-disease-virus-expressing-the-f-and-g-proteins-of-ampv-c-confers-protection-against-challenges-in-turkeys
#19
Haixia Hu, Jason P Roth, Laszlo Zsak, Qingzhong Yu
Avian metapneumovirus (AMPV) infects the respiratory and reproductive tracts of domestic poultry, resulting in substantial economic losses for producers. Live attenuated vaccines appear to be the most effective in countries where the disease is prevalent. However, reversion to virulence has been demonstrated in several studies. Therefore, the development of a stable and safe next generation vaccine against the AMPV disease is needed. In the present study, we generated a recombinant Newcastle disease virus (NDV) vectoring the fusion (F) protein and glycoprotein (G) genes of AMPV subtype-C (AMPV-C) as a bivalent vaccine candidate using reverse genetics technology...
June 22, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28642478/a-novel-plant-e3-ligase-stabilizes-escherichia-coli-heat-shock-factor-%C3%AF-32
#20
Yulong Niu, Xibing Xu, Chengcheng Liu, Tao Wang, Ke Liang, Jianmei Wang, Zhibin Liu, Xufeng Li, Yi Yang
The heat shock response is crucial for organisms against heat-damaged proteins and maintaining homeostasis at a high temperature. Heterologous expression of eukaryotic molecular chaperones protects Escherichia coli from heat stress. Here we report that expression of the plant E3 ligase BnTR1 significantly increases the thermotolerance of E. coli. Different from eukaryotic chaperones, BnTR1 expression induces the accumulation of heat shock factor σ(32) and heat shock proteins. The active site of BnTR1 in E...
June 22, 2017: Scientific Reports
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