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https://www.readbyqxmd.com/read/27906629/purification-activity-and-expression-levels-of-two-uridine-cytidine-kinase-isoforms-in-neuroblastoma-cell-lines
#1
R Meinsma, A B P van Kuilenburg
Uridine-cytidine kinase (UCK) catalyzes the phosphorylation of uridine, cytidine, and several pyrimidine ribonucleoside analogs. We overexpressed and purified the two known isoforms of human UCK in Escherichia coli, produced a specific antibody against UCK1 and characterized the kinetic properties of UCK1 and 2. The Vmax of purified recombinant UCK2 was 22- and 8-fold higher with uridine and cytidine, respectively, compared to those observed for the purified recombinant UCK1 enzyme. The Km of UCK1 was 39- and 40-fold higher with uridine and cytidine, respectively, compared to those observed for the purified recombinant UCK2 enzyme...
December 2016: Nucleosides, Nucleotides & Nucleic Acids
https://www.readbyqxmd.com/read/27902854/combining-affinity-chromatography-and-chromatofocusing-for-antibody-purification
#2
(no author information available yet)
No abstract text is available yet for this article.
January 2017: Biotechnology and Bioengineering
https://www.readbyqxmd.com/read/27902762/presenting-influenza-a-m2e-antigen-on-recombinant-spores-of-bacillus-subtilis
#3
Tomasz Łęga, Paulina Weiher, Michał Obuchowski, Dawid Nidzworski
Effective vaccination against influenza virus infection is a serious problem mainly due to antigenic variability of the virus. Among many of investigated antigens, the extracellular domain of the M2 protein (M2e) features high homology in all strains of influenza A viruses and antibodies against M2e and is protective in animal models; this makes it a potential candidate for generation of a universal influenza vaccine. However, due to the low immunogenicity of the M2e, formulation of a vaccine based on this antigen requires some modification to induce effective immune responses...
2016: PloS One
https://www.readbyqxmd.com/read/27894314/self-assembly-of-hexahistidine-tagged-tobacco-etch-virus-capsid-protein-into-microfilaments-that-induce-igg2-specific-response-against-a-soluble-porcine-reproductive-and-respiratory-syndrome-virus-chimeric-protein
#4
Carlos Alberto Manuel-Cabrera, Alba Adriana Vallejo-Cardona, Eduardo Padilla-Camberos, Rodolfo Hernández-Gutiérrez, Sara Elisa Herrera-Rodríguez, Abel Gutiérrez-Ortega
BACKGROUND: Assembly of recombinant capsid proteins into virus-like particles (VLPs) still represents an interesting challenge in virus-based nanotechnologies. The structure of VLPs has gained importance for the development and design of new adjuvants and antigen carriers. The potential of Tobacco etch virus capsid protein (TEV CP) as adjuvant has not been evaluated to date. FINDINGS: Two constructs for TEV CP expression in Escherichia coli were generated: a wild-type version (TEV-CP) and a C-terminal hexahistidine (His)-tagged version (His-TEV-CP)...
November 29, 2016: Virology Journal
https://www.readbyqxmd.com/read/27894306/first-phase-i-human-clinical-trial-of-a-killed-whole-hiv-1-vaccine-demonstration-of-its-safety-and-enhancement-of-anti-hiv-antibody-responses
#5
Eunsil Choi, Chad J Michalski, Seung Ho Choo, Gyoung Nyoun Kim, Elizabeth Banasikowska, Sangkyun Lee, Kunyu Wu, Hwa-Yong An, Anthony Mills, Stefan Schneider, U Fritz Bredeek, Daniel R Coulston, Shilei Ding, Andrés Finzi, Meijuan Tian, Katja Klein, Eric J Arts, Jamie F S Mann, Yong Gao, C Yong Kang
BACKGROUND: Vaccination with inactivated (killed) whole-virus particles has been used to prevent a wide range of viral diseases. However, for an HIV vaccine this approach has been largely negated due to inherent safety concerns, despite the ability of killed whole-virus vaccines to generate a strong, predominantly antibody-mediated immune response in vivo. HIV-1 Clade B NL4-3 was genetically modified by deleting the nef and vpu genes and substituting the coding sequence for the Env signal peptide with that of honeybee melittin signal peptide to produce a less virulent and more replication efficient virus...
November 28, 2016: Retrovirology
https://www.readbyqxmd.com/read/27890709/role-of-major-and-brain-specific-sgce-isoforms-in-the-pathogenesis-of-myoclonus-dystonia-syndrome
#6
Jianfeng Xiao, Satya R Vemula, Yi Xue, Mohammad M Khan, Francesca A Carlisle, Adrian J Waite, Derek J Blake, Ioannis Dragatsis, Yu Zhao, Mark S LeDoux
Loss-of-function mutations in SGCE, which encodes ε-sarcoglycan (ε-SG), cause myoclonus-dystonia syndrome (OMIM159900, DYT11). A "major" ε-SG protein derived from CCDS5637.1 (NM_003919.2) and a "brain-specific" protein, that includes sequence derived from alternative exon 11b (CCDS47642.1, NM_001099400.1), are reportedly localized in post- and pre-synaptic membrane fractions, respectively. Moreover, deficiency of the "brain-specific" isoform and other isoforms derived from exon 11b may be central to the pathogenesis of DYT11...
November 24, 2016: Neurobiology of Disease
https://www.readbyqxmd.com/read/27890399/a-partially-purified-outer-membrane-protein-virb9-1-for-low-cost-nanovaccines-against-anaplasma-marginale
#7
Liang Zhao, Antonino S Cavallaro, David Wibowo, Bing Zhang, Jun Zhang, Neena Mitter, Chengzhong Yu, Chun-Xia Zhao, Anton P J Middelberg
Anaplasma marginale is a devastating tick-borne pathogen causing anaplasmosis in cattle and results in significant economic loss to the cattle industry worldwide. Currently, there is no widely accepted vaccine against A. marginale. New generation subunit vaccines against A. marginale, which are much safer, more efficient and cost-effective, are in great need. The A. marginale outer membrane protein VirB9-1 is a promising antigen for vaccination. We previously have shown that soluble recombinant VirB9-1 protein can be expressed and purified from Escherichia coli and induce a high level of humoral and cellular immunity in mice...
November 24, 2016: Vaccine
https://www.readbyqxmd.com/read/27880036/protein-phosphotyrosine-proteome-profiling-by-superbinder-sh2-domain-affinity-purification-mass-spectrometry-ssh2-ap-ms
#8
Jiefei Tong, Biyin Cao, Gregory D Martyn, Jonathan R Krieger, Paul Taylor, Bradley Yates, Sachdev S Sidhu, Shawn S C Li, Xinliang Mao, Michael F Moran
Recently, "superbinder" SH2 domain variants with three amino acid substitutions (sSH2) were reported to have 100-fold or greater affinity for protein-phosphotyrosine (pY) than natural SH2 domains. Here we report a protocol in which His-tagged Src sSH2 efficiently captures pY-peptides from protease-digested HeLa cell total protein extracts. Affinity purification of pY-peptides by this method shows little bias for pY-proximal amino acid sequences, comparable to that achieved by using antibodies to pY, but with equal or higher yield...
November 23, 2016: Proteomics
https://www.readbyqxmd.com/read/27878624/the-creb-deubiquitinating-enzyme-does-not-directly-target-the-crea-repressor-protein-in-aspergillus-nidulans
#9
Md Ashiqul Alam, Niyom Kamlangdee, Joan M Kelly
Ubiquitination/deubiquitination pathways are now recognized as key components of gene regulatory mechanisms in eukaryotes. The major transcriptional repressor for carbon catabolite repression in Aspergillus nidulans is CreA, and mutational analysis led to the suggestion that a regulatory ubiquitination/deubiquitination pathway is involved. A key unanswered question is if and how this pathway, comprising CreB (deubiquitinating enzyme) and HulA (ubiquitin ligase) and other proteins, is involved in the regulatory mechanism...
November 23, 2016: Current Genetics
https://www.readbyqxmd.com/read/27871972/recombinant-antibody-fragment-production
#10
REVIEW
H Ma, R O'Kennedy
Recombinant antibodies are now very important in both therapeutics and diagnostics and offer significant advantages over conventional antibodies. The generation of a single-chain variable antibody fragment (scFv) (a common and important recombinant antibody format) is used to demonstrate the construction of a recombinant antibody library. An immunotube-based two-day panning approach, using Escherichia coli as an expression system, is utilised for antibody screening. The methods used for antibody selection and purification using immobilized metal affinity chromatography (IMAC) are described...
November 18, 2016: Methods: a Companion to Methods in Enzymology
https://www.readbyqxmd.com/read/27871373/design-and-production-of-antibodies-for-the-detection-of-streptococcus-uberis
#11
K Mihklepp, K Kivirand, M Nikopensius, D Peedel, M Utt, T Rinken
Streptococcus uberis (S. uberis) is an important environmental pathogen causing mastitis in dairy cattle. Mastitis or mammary gland infection is the most common disease in milking cows and cause significant economic burden to farmers due to the reduction of the amount of milk and its quality. The development of rapid analytical methods for the determination of mastitis-causing pathogens in milk is of utmost importance for the early identification of the causes of mastitis and the beginning of timely treatment of cows...
January 2017: Enzyme and Microbial Technology
https://www.readbyqxmd.com/read/27865853/the-principles-and-applications-of-avidin-based-nanoparticles-in-drug-delivery-and-diagnosis
#12
REVIEW
Akshay Jain, Kun Cheng
Avidin-biotin interaction is one of the strongest non-covalent interactions in the nature. Avidin and its analogues have therefore been extensively utilized as probes and affinity matrices for a wide variety of applications in biochemical assays, diagnosis, affinity purification, and drug delivery. Recently, there has been a growing interest in exploring this non-covalent interaction in nanoscale drug delivery systems for pharmaceutical agents, including small molecules, proteins, vaccines, monoclonal antibodies, and nucleic acids...
November 16, 2016: Journal of Controlled Release: Official Journal of the Controlled Release Society
https://www.readbyqxmd.com/read/27863521/seroprevalence-of-bovine-theileriosis-in-northern-china
#13
Yaqiong Li, Zhijie Liu, Junlong Liu, Jifei Yang, Qian Li, Pengfei Guo, Guiquan Guan, Guangyuan Liu, Jianxun Luo, Hong Yin, Youquan Li
BACKGROUND: Bovine theileriosis is a common disease transmitted by ticks, and can cause loss of beef and dairy cattle worldwide. Here, an indirect enzyme-linked immunosorbent assay (iELISA) based on Theileria luwenshuni surface protein (TlSP) was developed and used to carry out a seroepidemiological survey of bovine theileriosis in northern China. METHODS: We used the BugBuster Ni-NTA His•Bind Purification Kit to purify recombinant TlSP (rTlSP), which was subsequently analyzed by Western Blotting to evaluate cross-reactivity with other pathogen-positive sera...
November 18, 2016: Parasites & Vectors
https://www.readbyqxmd.com/read/27862786/efficient-conjugation-of-oligosaccharides-to-polymer-particles-by-furan-maleimide-diels-alder-reaction-application-to-the-capture-of-carbohydrate-binding-proteins
#14
Antoine Petrelli, Eric Samain, Stéphanie Pradeau, Sami Halila, Sebastien Fort
Glycan-protein interactions play a crucial role in physiological and pathological events. Hence, improving the isolation of carbohydrate-binding proteins (i.e. lectins and anti-glycan antibodies) from complex media can not only lead to a better understanding of their function, but also provides solutions for public health issues, such as water contamination or the need for universal blood plasma. Herein, we report a rapid and efficient method to produce carbohydrate-based affinity adsorbents combining enzymatic synthesis and metal-free click chemistry...
November 15, 2016: Chembiochem: a European Journal of Chemical Biology
https://www.readbyqxmd.com/read/27859689/protection-against-genital-tract-chlamydia-trachomatis-infection-following-intranasal-immunization-with-a-novel-recombinant-momp-vs2-4-antigen
#15
Ronza Hadad, Ellen Marks, Irina Kalbina, Karin Schön, Magnus Unemo, Nils Lycke, Åke Strid, Sören Andersson
The asymptomatic nature of most Chlamydia trachomatis infections and the lack of appropriate effects by current prevention and management call for vaccine development. We evaluated a recombinant subunit vaccine candidate based on the major outer membrane protein variable segments 2 and 4 (MOMP VS2/4). To achieve maximal immunogenicity and ease of production and purification, MOMP VS2/4 was constructed by using highly immunogenic sequences of MOMP only, thereby minimizing the presence of hydrophobic regions, and spacing the immunogenic epitopes with a flexible amino acid sequence...
December 2016: APMIS: Acta Pathologica, Microbiologica, et Immunologica Scandinavica
https://www.readbyqxmd.com/read/27855600/optimization-on-fc-for-improvement-of-stability-and-aggregation-resistance
#16
Xiaobo Chen, Fang Zeng, Tao Huang, Liang Cheng, Huan Liu, Rui Gong
Fc-based therapeutics including therapeutic full-size monoclonal antibodies (mAbs) and Fc-fusion proteins represent fastest-growing market in biopharmaceutical industrial. However, one major challenge during development of Fc-based therapeutics is how to maintain their efficacy in clinic use. Many factors may lead to failure in final marketing. For example, the stability and aggregation resistance might not be high enough for bearing the disadvantages during fermentation, purification, formulation, storage, shipment and other steps in manufacture and sale...
November 17, 2016: Current Pharmaceutical Biotechnology
https://www.readbyqxmd.com/read/27854147/improving-the-developability-of-an-anti-epha2-single-chain-variable-fragment-for-nanoparticle-targeting
#17
Melissa L Geddie, Neeraj Kohli, Dmitri B Kirpotin, Maja Razlog, Yang Jiao, Tad Kornaga, Rachel Rennard, Lihui Xu, Birgit Schoerberl, James D Marks, Daryl C Drummond, Alexey A Lugovskoy
Antibody-targeted nanoparticles have great promise as anti-cancer drugs; however, substantial developmental challenges of antibody modules prevent many candidates from reaching the clinic. Here, we describe a robust strategy for developing an EphA2-targeting antibody fragment for immunoliposomal drug delivery. A highly bioactive single-chain variable fragment (scFv) was engineered to overcome developmental liabilities, including low thermostability and weak binding to affinity purification resins. Improved thermostability was achieved by modifying the framework of the scFv, and complementarity-determining region (CDR)-H2 was modified to increase binding to protein A resins...
November 17, 2016: MAbs
https://www.readbyqxmd.com/read/27854020/production-and-purification-of-antibodies-against-histone-modifications
#18
Benoit Guillemette, Ian Hammond-Martel, Hugo Wurtele, Alain Verreault
Antibodies that recognize specific histone modifications are invaluable tools to study chromatin structure and function. There are numerous commercially available antibodies that recognize a remarkable diversity of histone modifications. Unfortunately, many of them fail to work in certain applications or lack the high degree of specificity required of these reagents. The production of affinity-purified polyclonal antibodies against histone modifications demands a little effort but, in return, provides extremely valuable tools that overcome many of the concerns and limitations of commercial antibodies...
2017: Methods in Molecular Biology
https://www.readbyqxmd.com/read/27853253/generation-of-metastatic-melanoma-specific-antibodies-by-affinity-purification
#19
Birgit Schütz, Anita Koppensteiner, David Schörghofer, Katharina Kinslechner, Gerald Timelthaler, Robert Eferl, Markus Hengstschläger, Albert Missbichler, Harald Hundsberger, Mario Mikula
Melanoma is the most aggressive type of skin cancer and one of the most frequent tumours in young adults. Identification of primary tumours prone to develop metastasis is of paramount importance for further patient stratification. However, till today, no markers exist that are routinely used to predict melanoma progression. To ameliorate this problem, we generated antiserum directed against metastatic melanoma tissue lysate and applied a novel approach to purify the obtained serum via consecutive affinity chromatography steps...
November 17, 2016: Scientific Reports
https://www.readbyqxmd.com/read/27852840/methyl-cytosine-based-immunoprecipitation-for-dna-methylation-analysis
#20
Paul M Lizardi, Qin Yan, Narendra Wajapeyee
In mammalian cells, DNA methylation at the 5-position of cytosine leads to recruitment of proteins that selectively recognize and bind 5-methylcytosine (5mC). Taking advantage of the structural identity of 5mC, various affinity purification-based protocols have been developed to enrich for either DNA that is modified by 5mC or proteins that recognize 5mC. In this protocol, an antibody against 5mC is used to immunoprecipitate the methylated DNA. The method can be scaled up to perform genome-wide DNA methylation analysis...
November 16, 2016: Cold Spring Harbor Protocols
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