A Single Mutation in Framework 2 of the Heavy Variable Domain Improves the Properties of a Diabody and a Related Single-Chain Antibody

Rodríguez-Rodríguez, Everardo Remi; Ledezma-Candanoza, Luis M.; Contreras-Ferrat, Luis Gabriel; Olamendi-Portugal, Timoteo; Possani, Lourival D.; Becerril, Baltazar; Riaño-Umbarila, Lidia

doi:10.1016/j.jmb.2012.07.007

Cited by 31 publications

(17 citation statements)

References 39 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Indeed, beneficial point mutations generated from independent maturation processes usually appear to be neutral or deleterious when combined [76]. A final variant (scFv LER) with increased thermal stability was created after a single point mutation of scFv LR in Framework 2 of the heavy chain capable of neutralizing 2 LD 50 of Cn2 toxin with no detectable symptoms when injected into mice after pre-incubation with the toxin in a 1/1 molar ratio [77]. …”

Section: Preclinical Evaluation Of Recombinant Antibodies For the mentioning

confidence: 99%

Engineering Venom’s Toxin-Neutralizing Antibody Fragments and Its Therapeutic Potential

Alvarenga

Zahid

Tommaso

et al. 2014

Toxins

View full text Add to dashboard Cite

Serum therapy remains the only specific treatment against envenoming, but anti-venoms are still prepared by fragmentation of polyclonal antibodies isolated from hyper-immunized horse serum. Most of these anti-venoms are considered to be efficient, but their production is tedious, and their use may be associated with adverse effects. Recombinant antibodies and smaller functional units are now emerging as credible alternatives and constitute a source of still unexploited biomolecules capable of neutralizing venoms. This review will be a walk through the technologies that have recently been applied leading to novel antibody formats with better properties in terms of homogeneity, specific activity and possible safety.

show abstract

Section: Preclinical Evaluation Of Recombinant Antibodies For the mentioning

confidence: 99%

Engineering Venom’s Toxin-Neutralizing Antibody Fragments and Its Therapeutic Potential

Alvarenga

Zahid

Tommaso

et al. 2014

Toxins

View full text Add to dashboard Cite

show abstract

“…This profile could be explained by new hydrogen bonds created between the alcohol functional group on tyrosine that interacts with the side chain of the glutamine at position 44. As previously described, VH-Q44 and VL-Q44 largely impact scFv stability as tyrosine VH-Y103 and VL-Y103 might also provide stronger VH/VL interaction [23,45]. To better understand the impact of VL 4 substitutions, M4V and P96Q mutations were introduced into the scFv S1B4 based on the better stability and production profiles observed for this molecule, rather than scFv S1A4, the latter having shown poorer features.…”

Section: Discussionmentioning

confidence: 99%

“…More precisely, additional work has been carried out on the structural role of framework regions (FRs). Several random mutation strategies have been undertaken to improve variable domains' stability [20,21] and/or the VH/VL interactions [22], stability, and activity [23,24]. Stability improvements and decrease in molecule aggregation were obtained by stabilizing the VH-βC"-βD loop using the VH-K64R substitution [25,26] or charged mutations [27][28][29][30].…”

Section: Introductionmentioning

confidence: 99%

Exploration and Modulation of Antibody Fragment Biophysical Properties by Replacing the Framework Region Sequences

et al. 2020

View full text Add to dashboard Cite

In order to increase the successful development of recombinant antibodies and fragments, it seems fundamental to enhance their expression and/or biophysical properties, such as the thermal, chemical, and pH stabilities. In this study, we employed a method bases on replacing the antibody framework region sequences, in order to promote more particularly single-chain Fragment variable (scFv) product quality. We provide evidence that mutations of the VH-C-C loop might significantly improve the prokaryote production of well-folded and functional fragments with a production yield multiplied by 27 times. Additional mutations are accountable for an increase in the thermal (+19.6 • C) and chemical (+1.9 M) stabilities have also been identified. Furthermore, the hereby-produced fragments have shown to remain stable at a pH of 2.0, which avoids molecule functional and structural impairments during the purification process. Lastly, this study provides relevant information to the understanding of the relationship between the antibodies amino acid sequences and their respective biophysical properties.

show abstract

“…While using a 5 residues linker, Rodriguez-Rodriguez et al demonstrated that a single mutation (E43G) in VH-FR2 of scFv56009F significantly improved the stability of the dimeric structure with positive consequences on its functional properties [24]. Future developments, directed evolution and fine 3-D structure analysis will help to understand these problems.…”

Section: Discussionmentioning

confidence: 99%

Generation of recombinant antibody fragments with toxin-neutralizing potential in loxoscelism

et al. 2016

View full text Add to dashboard Cite

Loxosceles spider bites often lead to serious envenomings and no definite therapy has yet been established. In such a context, it is of interest to consider an antibody-based targeted therapy. We have previously prepared a murine monoclonal IgG (LiMab7) that binds to 32-35kDa components of Loxosceles intermedia venom and neutralizes the dermonecrotic activity of the venom. Here, we re-engineered LiMab7 into a recombinant diabody. The protein was produced in bacteria and then it was functionally characterized. It proved to be efficient at neutralizing sphingomyelinase and hemolytic activities of the crude venom despite the slightly altered binding kinetic constants and the limited stability of the dimeric configuration. This is the first report of a specific recombinant antibody for a next-generation of Loxosceles antivenoms.

show abstract

A Single Mutation in Framework 2 of the Heavy Variable Domain Improves the Properties of a Diabody and a Related Single-Chain Antibody

Cited by 31 publications

References 39 publications

Engineering Venom’s Toxin-Neutralizing Antibody Fragments and Its Therapeutic Potential

Engineering Venom’s Toxin-Neutralizing Antibody Fragments and Its Therapeutic Potential

Exploration and Modulation of Antibody Fragment Biophysical Properties by Replacing the Framework Region Sequences

Generation of recombinant antibody fragments with toxin-neutralizing potential in loxoscelism

Contact Info

Product

Resources

About