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Protein engineering

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https://www.readbyqxmd.com/read/29792810/optogenetic-inhibition-of-g%C3%AE-q-protein-signaling-reduces-calcium-oscillation-stochasticity
#1
Pimkhuan Hannanta-Anan, Brian Y Chow
As fast terminators of G-protein coupled receptor (GPCR) signaling, regulators of G-protein signaling (RGS) serve critical roles in fine-tuning second messenger levels and, consequently, cellular responses to external stimuli. Here, we report the creation of an optogenetic RGS2 (opto-RGS2) that suppresses agonist-evoked calcium oscillations by the inactivation of Gαq protein. In this system, cryptochrome-mediated hetero-dimerization of the catalytic RGS2-box with its N-terminal amphipathic helix reconstitutes a functional membrane-localized complex that can dynamically suppress store-operated release of calcium...
May 24, 2018: ACS Synthetic Biology
https://www.readbyqxmd.com/read/29792795/tuning-the-morphology-of-nanostructured-peptide-films-by-introduction-of-a-secondary-structure-conformational-constraint-a-case-study-of-hierarchical-self-assembly
#2
Marta De Zotti, Beatrice Muzzi, Emanuela Gatto, Benedetta Di Napoli, Claudia Mazzuca, Antonio Palleschi, Ernesto Placidi, Fernando Formaggio, Claudio Toniolo, Mariano Venanzi
Peptide self-assembly is ubiquitous in nature. It governs the organization of proteins, controlling their folding kinetics and preserving their structure stability and bioactivity. In this connection, model oligopeptides may give important insights on the molecular mechanisms and elementary forces driving the formation of supramolecular structures. In this contribution we show that a single residue substitution, i.e. Aib (α-aminoisobutyric acid) in place of Ala at position 4 of an -(L-Ala)5- homo-oligomer, strongly alters the aggregation process...
May 24, 2018: Journal of Physical Chemistry. B
https://www.readbyqxmd.com/read/29792672/biosynthesis-of-lincosamide-antibiotics-reactions-associated-with-degradation-and-detoxification-pathways-play-a-constructive-role
#3
Daozhong Zhang, Zhijun Tang, Wen Liu
Natural products typically are small molecules produced by living organisms. These products possess a wide variety of biological activities and thus have historically played a critical role in medicinal chemistry and chemical biology either as chemotherapeutic agents or as useful tools. Natural products are not synthesized for use by human beings; rather, living organisms produce them in response to various biochemical processes and environmental concerns, both internal and external. These processes/concerns are often dynamic and thus motivate the diversification, optimization, and selection of small molecules in line with changes in biological function...
May 24, 2018: Accounts of Chemical Research
https://www.readbyqxmd.com/read/29792527/enteric-neural-stem-cells-expressing-insulin-like-growth-factor-1-a-novel-cellular-therapy-for-hirschsprung-s-disease-in-mouse-model
#4
Wei Liu, Lijuan Zhang, Rongde Wu
Transplanting enteric neural stem cells (ENSCs) is an innovative approach for replacing enteric neurons in Hirschsprung's disease (HSCR). However, posttransplantation cell survival and differentiation limit efficacy. We aimed to investigate whether transplantation of ENSCs engineered with insulin-like growth factor 1 (IGF-1) could improve survival and differentiation of the engrafted cells and promote functional recovery of the aganglionic colon. ENSCs were isolated from the intestine of neonatal mice and genetically modified to express IGF-1...
May 24, 2018: DNA and Cell Biology
https://www.readbyqxmd.com/read/29791796/tracking-of-engineered-bacteria-in-vivo-using-non-standard-amino-acid-incorporation
#5
Pichet Praveschotinunt, Noemie-Manuelle Dorval Courchesne, Ilona den Hartog, Chaochen Lu, Jessica J Kim, Peter Q Nguyen, Neel S Joshi
The rapidly growing field of microbiome research presents a need for better methods of monitoring gut microbes in vivo with high spatial and temporal resolution. We report a method of tracking microbes in vivo within the gastrointestinal tract by programming them to incorporate non-standard amino acids (NSAA) and labeling them via click chemistry. Using established machinery constituting an orthogonal translation system (OTS), we engineered Escherichia coli to incorporate p-azido-L-phenylalanine (pAzF) in place of the UAG (amber) stop codon...
May 23, 2018: ACS Synthetic Biology
https://www.readbyqxmd.com/read/29791150/elastin-like-polypeptide-elp-charge-influences-self-assembly-of-elp-mcherry-fusion-proteins
#6
Carolyn E Mills, Zachary Michaud, Bradley D Olsen
Self-assembly of protein-polymer bioconjugates presents an elegant strategy for controlling nanostructure and orientation of globular proteins in functional materials. Recent work has shown that genetic fusion of globular protein mCherry to an elastin-like polypeptide (ELP) yields similar self-assembly behavior to these protein-polymer bioconjugates. In the context of studying protein-polymer bioconjugate self-assembly, the mutability of the ELP sequence allows several different properties of the ELP block to be tuned orthogonally while maintaining consistent polypeptide backbone chemistry...
May 23, 2018: Biomacromolecules
https://www.readbyqxmd.com/read/29791133/design-construction-and-validation-of-histone-binding-effectors-in-vitro-and-in-cells
#7
Stefan J Tekel, Cassandra M Barrett, Daniel A Vargas, Karmella A Haynes
Chromatin is a system of nuclear proteins and nucleic acids that plays a pivotal role in gene expression and cell behavior, and is therefore the subject of intense study for cell development and cancer research. Biochemistry, crystallography, and reverse genetics have elucidated the macromolecular interactions that drive chromatin regulation. One of the central mechanisms is the recognition of post translational modifications (PTMs) on histone proteins by a family of nuclear proteins known as "readers...
May 23, 2018: Biochemistry
https://www.readbyqxmd.com/read/29789881/characterization-of-recombinant-e-coli-expressing-arsr-from-rhodopseudomonas-palustris-cga009-that-displays-highly-selective-arsenic-adsorption
#8
Changdong Ke, Chungui Zhao, Christopher Rensing, Suping Yang, Yi Zhang
Innovative methods to lower arsenic (As) exposure are sought. The As regulatory protein (ArsR) is reported of having high affinity and specificity to arsenite [As(III)]. Rhodopseudomonas palustris CGA009 is a good model organism for studying As detoxification due to at least three ars operons and four diverse arsRRP1-4 on the genome. In this study, four Escherichia coli harboring arsRRP1-4 derived from CGA009 were engineered and tested regarding their As resistance. The results showed that E. coli (arsRRP2 ) displayed robust As(III) resistance, and its growth inhibition rate was only 2...
May 22, 2018: Applied Microbiology and Biotechnology
https://www.readbyqxmd.com/read/29789544/development-of-muscular-dystrophy-in-a-crispr-engineered-mutant-rabbit-model-with-frame-disrupting-ano5-mutations
#9
Tingting Sui, Li Xu, Yeh Siang Lau, Di Liu, Tingjun Liu, Yandi Gao, Liangxue Lai, Renzhi Han, Zhanjun Li
Limb girdle muscular dystrophy type 2L (LGMD2L) and Miyoshi myopathy type 3 (MMD3) are autosomal recessive muscular dystrophy caused by mutations in the gene encoding anoctamin-5 (ANO5), which belongs to the anoctamin protein family. Two independent lines of mice with complete disruption of ANO5 transcripts did not exhibit overt muscular dystrophy phenotypes; instead, one of these mice was observed to present with some abnormality in sperm motility. In contrast, a third line of ANO5-knockout (KO) mice with residual expression of truncated ANO5 expression was reported to display defective membrane repair and very mild muscle pathology...
May 22, 2018: Cell Death & Disease
https://www.readbyqxmd.com/read/29789517/fluorescent-proteins-for-investigating-biological-events-in-acidic-environments
#10
REVIEW
Hajime Shinoda, Michael Shannon, Takeharu Nagai
The interior lumen of acidic organelles (e.g., endosomes, secretory granules, lysosomes and plant vacuoles) is an important platform for modification, transport and degradation of biomolecules as well as signal transduction, which remains challenging to investigate using conventional fluorescent proteins (FPs). Due to the highly acidic luminal environment (pH ~ 4.5⁻6.0), most FPs and related sensors are apt to lose their fluorescence. To address the need to image in acidic environments, several research groups have developed acid-tolerant FPs in a wide color range...
May 23, 2018: International Journal of Molecular Sciences
https://www.readbyqxmd.com/read/29789420/glycosylation-of-recombinant-anticancer-therapeutics-in-different-expression-systems-with-emerging-technologies
#11
REVIEW
Tariq Nadeem, Mohsin Ahmad Khan, Bushra Ijaz, Nadeem Ahmed, Zia Ur Rahman, Muhammad Shahzad Latif, Qurban Ali, Muhammad Adeel Rana
Glycosylation, a posttranslational modification, has a major role in recombinant anticancer therapeutic proteins, as most of the approved recombinant therapeutics are glycoproteins. The constant amino acid sequence of therapeutics determines the enzymatic activity, while the presence of glycans influences their pharmacokinetics, solubility, distribution, serum half-life, effector function, and binding to receptors. Glycoproteins expressed in different expression systems acquire their own oligosaccharides, which increases the protein diversity...
May 22, 2018: Cancer Research
https://www.readbyqxmd.com/read/29789067/engineering-aspects-of-protein-interactions-and-self-assembly
#12
Miriam Linsenmeier, Paolo Arosio
In the new Laboratory for Biochemical Engineering (LBCE) at ETH Zurich researchers combine principles of chemical engineering with microfluidic technology and biophysical methods to investigate the physical determinants of biomolecular self-assembly in living organisms. In this account, we show the impact of this activity on concrete applications in biomedical sciences and biotechnology. We focus in particular on the field of protein aggregation and phase separation, and we highlight examples in the context of diagnosis and treatment of Alzheimer's disease and neurodegenerative disorders, cell compartmentalization as well as manufacturing and delivery of therapeutic proteins...
May 30, 2018: Chimia
https://www.readbyqxmd.com/read/29788485/enzymatic-synthesis-of-random-sequences-of-rna-and-rna-analogues-by-dna-polymerase-theta-mutants-for-the-generation-of-aptamer-libraries
#13
Irina Randrianjatovo-Gbalou, Sandrine Rosario, Odile Sismeiro, Hugo Varet, Rachel Legendre, Jean-Yves Coppée, Valérie Huteau, Sylvie Pochet, Marc Delarue
Nucleic acid aptamers, especially RNA, exhibit valuable advantages compared to protein therapeutics in terms of size, affinity and specificity. However, the synthesis of libraries of large random RNAs is still difficult and expensive. The engineering of polymerases able to directly generate these libraries has the potential to replace the chemical synthesis approach. Here, we start with a DNA polymerase that already displays a significant template-free nucleotidyltransferase activity, human DNA polymerase theta, and we mutate it based on the knowledge of its three-dimensional structure as well as previous mutational studies on members of the same polA family...
May 21, 2018: Nucleic Acids Research
https://www.readbyqxmd.com/read/29788407/deregulation-of-sgk1-in-ulcerative-colitis-a-paradoxical-relationship-between-immune-cells-and-colonic-epithelial-cells
#14
Rocco Spagnuolo, Vincenzo Dattilo, Lucia D'Antona, Cristina Cosco, Rossana Tallerico, Valeria Ventura, Francesco Conforti, Caterina Camastra, Rosellina M Mancina, Giada Catalogna, Vincenzo Cosco, Rodolfo Iuliano, Ennio Carbone, Nicola Perrotti, Rosario Amato, Patrizia Doldo
Background: Inflammatory bowel disease (IBD) is due to the interaction of genetic and environmental factors that trigger an unbalanced immune response ultimately resulting in the peculiar inflammatory reaction. Experimental models of IBD point to a role of T-cell-derived cytokines (Th17) and to SGK1 as mediator of the Th17 switch. We hypothesize that SGK1, a salt inducible kinase, directs lymphocytic behavior and tissue damage. Methods: Eleven controls and 32 ulcerative colitis (UC) patients were randomized according to endoscopic Mayo score...
May 18, 2018: Inflammatory Bowel Diseases
https://www.readbyqxmd.com/read/29788355/swiss-model-homology-modelling-of-protein-structures-and-complexes
#15
Andrew Waterhouse, Martino Bertoni, Stefan Bienert, Gabriel Studer, Gerardo Tauriello, Rafal Gumienny, Florian T Heer, Tjaart A P de Beer, Christine Rempfer, Lorenza Bordoli, Rosalba Lepore, Torsten Schwede
Homology modelling has matured into an important technique in structural biology, significantly contributing to narrowing the gap between known protein sequences and experimentally determined structures. Fully automated workflows and servers simplify and streamline the homology modelling process, also allowing users without a specific computational expertise to generate reliable protein models and have easy access to modelling results, their visualization and interpretation. Here, we present an update to the SWISS-MODEL server, which pioneered the field of automated modelling 25 years ago and been continuously further developed...
May 21, 2018: Nucleic Acids Research
https://www.readbyqxmd.com/read/29787277/targeting-of-egfr-vegfr2-and-akt-by-engineered-dual-drug-encapsulated-mesoporous-silica-gold-nanoclusters-sensitizes-tamoxifen-resistant-breast-cancer
#16
Bn Prashanth Kumar, Nagaprasad Puvvada, Shashi Rajput, Siddik Sarkar, Madhusudan Kr Mahto, Murali M Yallapu, Amita Pathak, Luni Emdad, Swadesh K Das, Rui L Reis, Subhas C Kundu, Paul B Fisher, Mahitosh Mandal
Tamoxifen has improved overall disease-free survival and reduced mortality rates in cancer patients. However, breast cancer patients often fail to respond to tamoxifen therapy due to the development of a drug-resistant phenotype. Functional analysis and molecular studies suggest that protein mutation and dysregulation of survival signaling molecules such as epidermal growth factor receptor, vascular endothelial growth factor receptor 2 and Akt contribute to tamoxifen resistance. Various strategies, including combinatorial therapies, show to chemo-sensitize tamoxifen-resistant cancers...
May 22, 2018: Molecular Pharmaceutics
https://www.readbyqxmd.com/read/29787234/unfolding-the-mysteries-of-protein-metamorphosis
#17
Acacia F Dishman, Brian F Volkman
Since the proposal of Anfinsen's thermodynamic hypothesis in 1963, our understanding of protein folding and dynamics has gained significant appreciation of its nuance and complexity. Intrinsically disordered proteins, chameleonic sequences, morpheeins, and metamorphic proteins have broadened the protein folding paradigm. Here, we discuss noncanonical protein folding patterns, with an emphasis on metamorphic proteins, and we review known metamorphic proteins that occur naturally and that have been engineered in the laboratory...
May 22, 2018: ACS Chemical Biology
https://www.readbyqxmd.com/read/29786902/engineered-metalloenzymes-with-non-canonical-coordination-environments
#18
Takahiro Hayashi, Donald Hilvert, Anthony P Green
Nature employs a limited number of genetically encoded, metal coordinating residues to create metalloenzymes with diverse structures and functions. Engineered components of the cellular translation machinery can now be exploited to genetically encode non-canonical ligands with user-defined electronic and structural properties. This ability to efficiently install 'chemically programmed' ligands into proteins can provide powerful chemical probes of metalloenzyme mechanisms and presents excellent opportunities to create metalloprotein catalysts with augmented properties and novel activities...
May 22, 2018: Chemistry: a European Journal
https://www.readbyqxmd.com/read/29786899/improved-stable-indocyanine-green-icg-mediated-cancer-optotheranostics-with-naturalized-hepatitis-b-core-particles
#19
Wenjun Shan, Ronghe Chen, Qiang Zhang, Jie Zhao, Binbin Chen, Xi Zhou, Shefang Ye, Shengli Bi, Liming Nie, Lei Ren
In recent years, hepatitis B core protein virus-like particle (HBc VLP) is an impressive biomaterial, which has attracted considerable attention due to favorable properties such as structural stability, high uptake efficiency, and biocompatibility in biomedical applications. Heretofore, only a few attempts have been made to apply it in physical, chemical, and biological therapy for cancer. In this study, a tumor-targeting RGD-HBc VLP is first fabricated through genetic engineering. For image-guided cancer phototherapy, indocyanine green (ICG) is loaded into RGD-HBc VLP via a disassembly/reassembly pathway and electrostatic attraction with high efficiency...
May 22, 2018: Advanced Materials
https://www.readbyqxmd.com/read/29786560/microfabrication-of-liver-and-heart-tissues-for-drug-development
#20
REVIEW
Grace E Brown, Salman R Khetani
Drug-induced liver- and cardiotoxicity remain among the leading causes of preclinical and clinical drug attrition, marketplace drug withdrawals and black-box warnings on marketed drugs. Unfortunately, animal testing has proven to be insufficient for accurately predicting drug-induced liver- and cardiotoxicity across many drug classes, likely due to significant differences in tissue functions across species. Thus, the field of in vitro human tissue engineering has gained increasing importance over the last 10 years...
July 5, 2018: Philosophical Transactions of the Royal Society of London. Series B, Biological Sciences
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