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Order Picornavirales
Published in Paul Pumpens, Peter Pushko, Philippe Le Mercier, Virus-Like Particles, 2022
Paul Pumpens, Peter Pushko, Philippe Le Mercier
Koho et al. (2015) presented a nanocarrier platform based on the modified NoV VLPs and noncovalent chemical conjugation. The VLPs were modified by adding a C-terminal polyhistidine tag, which projected out of the VLP surface, produced by the baculovirus expression system. The norovirus genotype GII.4 utilized in this study has been previously expressed as unmodified VLP by Allen et al. (2009) and Koho et al. (2012). The chimeric NoV VLPs were generated by only the His-tagged capsid proteins or in the form of mosaic particles containing both the His-tagged and the original wild-type NoV capsid protein. The polyhistidine tag was first utilized in VLP purification and later employed to attach different cargo molecules noncovalently on the VLP surface via tris-nitrilotriacetic acid (trisNTA) adaptors. The surface-modified NoV VLP nanocarriers were implemented first in delivering a conjugated fluorescent dye as a model molecule into human cells. Generally, this technology provided a universal nanoparticle tool for the adaptation of vaccine delivery or targeting, for increasing and fine-tuning vaccine immunogenicity or bioavailability via a displayable molecule, and for use as surface building blocks (Koho et al. 2015).
Aptamers and Cancer Nanotechnology
Published in Mansoor M. Amiji, Nanotechnology for Cancer Therapy, 2006
Omid C. Farokhzad, Sangyong Jon, Robert Langer
Covalent conjugation of aptamers to substrates or drug delivery vehicles can be achieved most commonly through succinimidyl ester–amine chemistry that results in a stable amide linkage43,43 or through maleimide–thiol chemistry.23,108,116–120 Potential noncovalent strategies include affinity interactions (i.e., streptavidin–biotin) and metal coordination (i.e., between a polyhistidine tag at the end of the aptamer and Ni+2 chelates with immobilized nitrilotriacetic acid on the surface of the polymer particles). These covalent and noncovalent strategies have been used to immobilize a wide range of biomolecules, including proteins, enzymes, peptides, and nucleic acids to delivery vehicles.
Zika virus pathogenesis and current therapeutic advances
Published in Pathogens and Global Health, 2021
Caroline Mwaliko, Raphael Nyaruaba, Lu Zhao, Evans Atoni, Samuel Karungu, Matilu Mwau, Dimitri Lavillette, Han Xia, Zhiming Yuan
Kim et al. designed a recombinant E1/E3-deleted adenoviral vector (pAd.ZIKV-Efl) from the ZIKV strain BeH815744 that expressed a codon-optimized Env-antigen. The extracellular region of the ZIKV envelope was linked to the T4 fibrin trimerization domain, facilitating protein expression. Additionally, the vector was designed with a polyhistidine tag and a Tobacco Etch Virus (TEV) to increase protein folding and facilitate purification. The vaccine was shown to protect C57BL/6 mice from lethal challenge with the ZIKV DAKAR41542 strain [100]. Notably, in these vaccine studies, the ZIKV E subunit protein production yield was very low, a finding that has also been highlighted by Larroca et al. [93]. The low yield of the E protein was probably due to the absence of preM, which is important for protein stability. Therefore, preM is an indispensable factor in the development of ZIKV E protein-based vaccines.
Productive common light chain libraries yield diverse panels of high affinity bispecific antibodies
Published in mAbs, 2018
Thomas Van Blarcom, Kevin Lindquist, Zea Melton, Wai Ling Cheung, Chris Wagstrom, Dan McDonough, Cendy Valle Oseguera, Sheng Ding, Andrea Rossi, Shobha Potluri, Purnima Sundar, Steven Pitts, Marina Sirota, Meri Galindo Casas, Yu Yan, Jeffrey Jones, Zygy Roe-Zurz, Surabhi Srivatsa Srinivasan, Wenwu Zhai, Jaume Pons, Arvind Rajpal, Javier Chaparro-Riggers
Two model antibodies that utilize the same common light chain, but bind to different antigens were cloned into either the hIgG2 RRRR construct with up to a 10x polyhistidine tag or the hIgG2 EEE construct, then expressed individually or co-expressed in mammalian cells. The clarified cell culture supernatants were purified via fast protein liquid chromatography (FPLC) using Protein A and IEC. The IEC fractions expected to be the BsIgG were collected and dialyzed in phosphate-buffered saline, then confirmed to be the BsIgG by antigen binding using bio-layer interferometry (BLI). As expected, the separation of the BsIgG from the monospecific contaminants increased proportionally to the length of the polyhistidine tag, leading to robust separation of the BsIgG from the monospecific contaminants (Fig. 4B). While the 10x polyhistidine tag resulted in the most robust separation, the inclusion of the polyhistidine tag raised concerns that it may increase non-specific binding. This concern was addressed by incubating each BsIgG at up to 100 nM with cells that do not express either target antigen, and then detecting bound IgG with anti-human IgG Alexa Fluor 647 by flow cytometry. Only the BsIgG with the 10x polyhistidine tag showed evidence of non-specific binding (Fig. S13).
Anion inhibition studies of a beta carbonic anhydrase from the malaria mosquito Anopheles gambiae
Published in Journal of Enzyme Inhibition and Medicinal Chemistry, 2018
Daniela Vullo, Leo Syrjänen, Marianne Kuuslahti, Seppo Parkkila, Claudiu T. Supuran
For recombinant protein production, the β-CA gene was constructed and cloned into the pFastBac1TM vector. The forward primer used in the initial amplification of the β-CA gene was 5′-CGCGGATCCATGGAGCGTATATTGCGAGGC-3′ (F2), and the reverse primer was 5′-GCCCTCGAGTTAATGGTGGTGATGGTGGTGGGAACC-ACGGGGCACCAGCGAATAGTATCGCCGTACCTC-3′ (R2). The latter primer contains nucleotide repeats to create the C-terminal polyhistidine tag with six histidines. In addition, the forward primer contained the restriction site for BamHI and the reverse primer for XhoI. The reverse primer also contained the nucleotide sequence encoding thrombin cleavage site. The PCR program was as follows: 98 °C for 30 s; then 35 cycles of 98 °C for 10 s, 62 °C for 15 s, and 62 °C for 30 s, and finally 72 °C for 5 min.