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Protein engineering

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https://www.readbyqxmd.com/read/28917797/a-rapid-and-specific-colorimetric-method-for-free-tryptophan-quantification
#1
Yinan Wu, Tianmin Wang, Chong Zhang, Xin-Hui Xing
Tryptophan is one of the eight essential amino acids and plays an important role in many biological processes. For its interaction with human health, environment and relevant commercial interest in biotechnology-based production, rapid and specific quantification method for this molecule accessible to common laboratories is badly needed. We herein reported a simple colorimetric method for free tryptophan quantification with 96-well-plate-level throughput. Our protocol firstly converted tryptophan to indole enzymatically by purified tryptophanases and then used reactivity of indole with hydroxylamine to form pink product with absorption peak at 530nm, enabling the quantification of tryptophan with simple spectrometry in just two hours...
January 1, 2018: Talanta
https://www.readbyqxmd.com/read/28917741/immobilization-of-cytochrome-c-and-its-application-as-electrochemical-biosensors
#2
REVIEW
Zahra Sadat Aghamiri, Mohsen Mohsennia, Hossain-Ali Rafiee-Pour
Cytochrome c (Cyt c) has been used as a model protein to investigate the characters of modified electrodes by many researchers. It has been also employed to construct biosensors to detect hydrogen peroxide, nitrate, superoxide, and etc. Cyt c immobilization techniques, including physical adsorption, entrapment in hydrogel or polymers, layer-by-layer assembly, Langmuir-Blodgett, and covalent attachment are discussed followed by various electrochemical methods applied in the electrode modification. The exploration of some modified protein electrodes, for example, screen printed, microperoxidase and engineered Cyt c are also presented...
January 1, 2018: Talanta
https://www.readbyqxmd.com/read/28916751/understanding-cry2-interactions-for-optical-control-of-intracellular-signaling
#3
Liting Duan, Jen Hope, Qunxiang Ong, Hsin-Ya Lou, Namdoo Kim, Comfrey McCarthy, Victor Acero, Michael Z Lin, Bianxiao Cui
Arabidopsis cryptochrome 2 (CRY2) can simultaneously undergo light-dependent CRY2-CRY2 homo-oligomerization and CRY2-CIB1 hetero-dimerization, both of which have been widely used to optically control intracellular processes. Applications using CRY2-CIB1 interaction desire minimal CRY2 homo-oligomerization to avoid unintended complications, while those utilizing CRY2-CRY2 interaction prefer robust homo-oligomerization. However, selecting the type of CRY2 interaction has not been possible as the molecular mechanisms underlying CRY2 interactions are unknown...
September 15, 2017: Nature Communications
https://www.readbyqxmd.com/read/28916265/glycans-function-as-anchors-for-antibodies-and-help-drive-hiv-broadly-neutralizing-antibody-development
#4
Raiees Andrabi, Ching-Yao Su, Chi-Hui Liang, Sachin S Shivatare, Bryan Briney, James E Voss, Salar Khan Nawazi, Chung-Yi Wu, Chi-Huey Wong, Dennis R Burton
Apex broadly neutralizing HIV antibodies (bnAbs) recognize glycans and protein surface close to the 3-fold axis of the envelope (Env) trimer and are among the most potent and broad Abs described. The evolution of apex bnAbs from one donor (CAP256) has been studied in detail and many Abs at different stages of maturation have been described. Using diverse engineering tools, we investigated the involvement of glycan recognition in the development of the CAP256.VRC26 Ab lineage. We found that sialic acid-bearing glycans were recognized by germline-encoded and somatically mutated residues on the Ab heavy chain...
September 8, 2017: Immunity
https://www.readbyqxmd.com/read/28916111/icosahedral-plant-viral-nanoparticles-bioinspired-synthesis-of-nanomaterials-nanostructures
#5
REVIEW
Kannan Badri Narayanan, Sung Soo Han
Viral nanotechnology utilizes virus nanoparticles (VNPs) and virus-like nanoparticles (VLPs) of plant viruses as highly versatile platforms for materials synthesis and molecular entrapment that can be used in the nanotechnological fields, such as in next-generation nanoelectronics, nanocatalysis, biosensing and optics, and biomedical applications, such as for targeting, therapeutic delivery, and non-invasive in vivo imaging with high specificity and selectivity. In particular, plant virus capsids provide biotemplates for the production of novel nanostructured materials with organic/inorganic moieties incorporated in a very precise and controlled manner...
August 31, 2017: Advances in Colloid and Interface Science
https://www.readbyqxmd.com/read/28915385/photoluminescent-lateral-flow-based-on-non-radiative-energy-transfer-for-protein-detection-in-human-serum
#6
Alejandro Zamora-Gálvez, Eden Morales-Narváez, Javier Romero, Arben Merkoçi
A new paper-based lateral flow immunoassay configuration was engineered and investigated. The assay is intended for the detection of a model protein in human serum, that is, human immunoglobulin G, with the aim to demonstrate a virtually universal protein detection platform. Once the sample is added in the strip, the analyte is selectively captured by antibody-decorated silica beads (Ab-SiO2) onto the conjugate pad and the sample flows by capillarity throughout the strip until reaching the test line, where a sandwich-like immunocomplex takes place due to the presence of antibody-functionalized QDs (Ab-QDs) onto the test line...
September 11, 2017: Biosensors & Bioelectronics
https://www.readbyqxmd.com/read/28915034/affinity-guided-oxime-chemistry-for-selective-protein-acylation-in-live-tissue-systems
#7
Tomonori Tamura, Zhining Song, Kazuma Amaike, Shin Lee, Sifei Yin, Shigeki Kiyonaka, Itaru Hamachi
Catalyst-mediated protein modification is a powerful approach for the imaging and engineering of natural proteins. We have previously developed affinity-guided 4-dimethylaminopyridine (DMAP) (AGD) chemistry as an efficient protein modification method using a catalytic acyl transfer reaction. However, because of the high electrophilicity of the thioester acyl donor molecule, AGD chemistry suffers from nonspecific reactions to proteins other than the target protein in crude biological environments, such as cell lysates, live cells, and tissue samples...
September 15, 2017: Journal of the American Chemical Society
https://www.readbyqxmd.com/read/28914771/geminiviruses-and-plant-hosts-a-closer-examination-of-the-molecular-arms-race
#8
REVIEW
Shunmugiah V Ramesh, Pranav P Sahu, Manoj Prasad, Shelly Praveen, Hanu R Pappu
Geminiviruses are plant-infecting viruses characterized by a single-stranded DNA (ssDNA) genome. Geminivirus-derived proteins are multifunctional and effective regulators in modulating the host cellular processes resulting in successful infection. Virus-host interactions result in changes in host gene expression patterns, reprogram plant signaling controls, disrupt central cellular metabolic pathways, impair plant's defense system, and effectively evade RNA silencing response leading to host susceptibility...
September 15, 2017: Viruses
https://www.readbyqxmd.com/read/28914478/mechanistic-insights-into-cyclic-peptide-generation-by-dnae-split-inteins-through-quantitative-and-structural-investigation
#9
Leonhard Kick, Sabrina Harteis, Maximilian F Koch, Sabine Schneider
Inteins carry out protein-splicing reactions, which are utilised in protein chemistry, protein engineering and biotechnological applications. Rearrangement of the order of the domains in split-inteins results in a head-to-tail cyclisation of the target sequence, which can be used to genetically encode and express libraries of cyclic peptides. The efficiency of the splicing reaction depends on the target sequence. Here we used mass spectrometry to assess the cyclic peptide formation of different, hexameric target sequences in vivo by the DnaE split-inteins from Synechocystis sp...
September 15, 2017: Chembiochem: a European Journal of Chemical Biology
https://www.readbyqxmd.com/read/28913796/allergenicity-assessment-of-transgenic-wheat-lines-in-silico
#10
Ankita Mishra, Naveen Arora
Agriculture biotechnology is a promising tool for developing varieties with enhanced quality and quantity. Transgenic proteins expressed by genetically modified (GM) food crops improve crop characteristics like nutritional value, taste, and texture, and endow plants with resistance against fungus, pests, and insects. Despite such potential benefits, there are concerns regarding possible adverse effects of GM crops on human health, animals and the environment. Among the proposed guidelines for GM food safety testing-the weight-of-evidence approach proposed by the Codex Alimentarius Commission (ALINORM 03/34A) is the most recent...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/28913776/interactions-of-antisera-to-different-chlamydia-and-chlamydophila-species-with-the-ribosomal-protein-rps27a-correlate-with-impaired-protein-synthesis-in-a-human-choroid-plexus-papilloma-cell-line
#11
Abdullah Almamy, Christian Schwerk, Horst Schroten, Hiroshi Ishikawa, Abdul Rahman Asif, Bernhard Reuss
Chlamydia trachomatis (CT) and the Chlamydophila species (CS) Chlamydophila pneumoniae (CPn), and Chlamydophila psittaci (CPs) are suggested to induce autoantibodies causative of several human autoimmune disorders like rheumatoid arthritis and systemic lupus erythematosus (SLE). The aim of the present study was therefore to identify cellular protein interaction partners with antisera to CT (α-CT) or CS (α-CS) and to identify functional consequences of such interaction in vitro. As detected with a commercial first trimester human prenatal brain multiprotein array (hEXselect, Engine, Germany), the most frequent interaction partner with both α-CT and α-CS was the ribosomal small subunit protein RPS27a...
September 14, 2017: Immunologic Research
https://www.readbyqxmd.com/read/28913758/a-review-of-hydrogen-production-by-photosynthetic-organisms-using-whole-cell-and-cell-free-systems
#12
Baker A Martin, Paul D Frymier
Molecular hydrogen is a promising currency in the future energy economy due to the uncertain availability of finite fossil fuel resources and environmental effects from their combustion. It also has important uses in the production of fertilizers and platform chemicals as well as in upgrading conventional fuels. Conventional methods for producing molecular hydrogen from natural gas produce carbon dioxide and use a finite resource as feedstock. However, these issues can be overcome by using light energy from the Sun combined with microorganisms and their molecular machinery capable of photosynthesis...
September 14, 2017: Applied Biochemistry and Biotechnology
https://www.readbyqxmd.com/read/28912946/crosslinking-method-of-hyaluronic-based-hydrogel-for-biomedical-applications
#13
Sureerat Khunmanee, Younghyen Jeong, Hansoo Park
In the field of tissue engineering, there is a need for advancement beyond conventional scaffolds and preformed hydrogels. Injectable hydrogels have gained wider admiration among researchers as they can be used in minimally invasive surgical procedures. Injectable gels completely fill the defect area and have good permeability and hence are promising biomaterials. The technique can be effectively applied to deliver a wide range of bioactive agents, such as drugs, proteins, growth factors, and even living cells...
January 2017: Journal of Tissue Engineering
https://www.readbyqxmd.com/read/28912594/quantification-of-aquaporin-z-reconstituted-into-vesicles-for-biomimetic-membrane-fabrication
#14
Hui Xian Gan, Hu Zhou, Qingsong Lin, Yen Wah Tong
Aquaporin incorporated biomimetic membranes are anticipated to offer unprecedented desalination capabilities. However, the lack of accurate methods to quantify the reconstituted aquaporin presents a huge hurdle in investigating aquaporin performance and optimizing membrane fabrication. Herein, we present three quantification methods to determine the Aquaporin-Z reconstituted into E. coli lipid vesicles: 1) nanogold labeling with transmission electron microscopy (TEM) visualization, 2) nickel labeling with inductively coupled plasma-mass spectrometry (ICP-MS) and 3) gel electrophoresis...
September 14, 2017: Scientific Reports
https://www.readbyqxmd.com/read/28911198/loss-of-native-%C3%AE-synuclein-multimerization-by-strategically-mutating-its-amphipathic-helix-causes-abnormal-vesicle-interactions-in-neuronal-cells
#15
Ulf Dettmer, Nagendran Ramalingam, Victoria E von Saucken, Tae-Eun Kim, Andrew J Newman, Elizabeth Terry-Kantor, Silke Nuber, Maria Ericsson, Saranna Fanning, Tim Bartels, Susan Lindquist, Oren A Levy, Dennis Selkoe
α-Synuclein (αS) forms round cytoplasmic inclusions in Parkinson's disease (PD) and dementia with Lewy bodies (DLB). Evidence suggests a physiological function of αS in vesicle trafficking and release. In contrast to earlier tenets, recent work indicates that αS normally exists in cells in a dynamic equilibrium between monomers and tetramers/multimers. We engineered αS mutants incapable of multimerization, leading to excess monomers at vesicle membranes. By EM, such mutants induced prominent vesicle clustering, leading to round cytoplasmic inclusions...
September 15, 2017: Human Molecular Genetics
https://www.readbyqxmd.com/read/28911185/identification-of-a-new-gene-yecc-involved-in-threonine-export-in-escherichia-coli
#16
Yiran Xu, Yongfei Liu, Feiran Li, Guoqiang Cao, Ping Zheng, Jibin Sun, Jianping Wen, Dawei Zhang
Threonine (Thr), an essential amino acid for mammals, has expanded its application to industries with fast-growing market demand. One factor that limits the production of L-threonine by microbial fermentation is the lack of an effective export system. This study proposes a method based on isobaric tags for relative and absolute quantification (iTRAQ) to determine potential Thr excretion proteins. Proteomics analysis revealed that an ABC family transport protein named YecC was upregulated in response to Thr addition...
September 15, 2017: FEMS Microbiology Letters
https://www.readbyqxmd.com/read/28911121/engineering-human-primpol-into-an-efficient-rna-dependent-dna-primase-polymerase
#17
Rubén Agudo, Patricia A Calvo, María I Martínez-Jiménez, Luis Blanco
We have developed a straightforward fluorometric assay to measure primase-polymerase activity of human PrimPol (HsPrimPol). The sensitivity of this procedure uncovered a novel RNA-dependent DNA priming-polymerization activity (RdDP) of this enzyme. In an attempt to enhance HsPrimPol RdDP activity, we constructed a smart mutant library guided by prior sequence-function analysis, and tested this library in an adapted screening platform of our fluorometric assay. After screening less than 500 variants, we found a specific HsPrimPol mutant, Y89R, which displays 10-fold higher RdDP activity than the wild-type enzyme...
September 6, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28911120/phirex-a-programmable-and-red-light-regulated-protein-expression-switch-for-yeast
#18
Lena Hochrein, Fabian Machens, Katrin Messerschmidt, Bernd Mueller-Roeber
Highly regulated induction systems enabling dose-dependent and reversible fine-tuning of protein expression output are beneficial for engineering complex biosynthetic pathways. To address this, we developed PhiReX, a novel red/far-red light-regulated protein expression system for use in Saccharomyces cerevisiae. PhiReX is based on the combination of a customizable synTALE DNA-binding domain, the VP64 activation domain and the light-sensitive dimerization of the photoreceptor PhyB and its interacting partner PIF3 from Arabidopsis thaliana...
September 6, 2017: Nucleic Acids Research
https://www.readbyqxmd.com/read/28905106/recombinant-mouse-osteocalcin-secreted-by-lactococcus-lactis-promotes-glucagon-like-peptide-1-induction-in-stc-1-cells
#19
Fu Namai, Suguru Shigemori, Koichi Sudo, Takashi Sato, Yoshinari Yamamoto, Shireen Nigar, Tasuku Ogita, Takeshi Shimosato
An osteoblastic protein, osteocalcin (OC), exists in vivo in two forms: carboxylated OC, and uncarboxylated or low-carboxylated OC (ucOC). ucOC acts as a hormone to regulate carbon and energy metabolism. Recent studies demonstrated that ucOC exerts insulinotropic effects, mainly through the glucagon-like peptide 1 (GLP-1) pathway. GLP-1 is an insulinotropic hormone secreted by enteroendocrine L cells in the small intestine. Thus, efficient delivery of ucOC to the small intestine may be a new therapeutic option for metabolic diseases such as diabetes and obesity...
September 13, 2017: Current Microbiology
https://www.readbyqxmd.com/read/28904863/chimeric-zhhh-neuroglobin-acts-as-a-cell-membrane-penetrating-inducer-of-neurite-outgrowth
#20
Nozomu Takahashi, Wataru Onozuka, Seiji Watanabe, Keisuke Wakasugi
Neuroglobin (Ngb) is a heme protein expressed in the vertebrate brain. We previously engineered a chimeric Ngb protein, in which module M1 of human Ngb is replaced by that of zebrafish Ngb, and showed that the chimeric ZHHH Ngb has a cell membrane-penetrating activity similar to that of zebrafish Ngb and also rescues cells from death caused by hypoxia/reoxygenation as does human Ngb. Recently, it was reported that overexpression of mammalian Ngb in neuronal cells induces neurite outgrowth. In this study, we performed neurite outgrowth assays of chimeric Ngb using rat pheochromocytoma PC12 cells...
September 2017: FEBS Open Bio
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