scholarly journals Nanoscale spatial dependence of avidity in an IgG1 antibody

2019 ◽  
Author(s):  
Agnieszka Jendroszek ◽  
Magnus Kjaergaard
Keyword(s):  
Immune System ◽  
Spatial Dependence ◽  
Target Recognition ◽  
Coiled Coil ◽  
Coiled Coils ◽  
Antibody Engineering ◽  
Bispecific Antibodies ◽  
Secreted Proteins ◽  
Biophysical Techniques ◽  
The Stability

AbstractAntibodies are secreted proteins that are crucial to recognition of pathogens by the immune system and are also efficient pharmaceuticals. The affinity and specificity of target recognition can increase remarkably through avidity effects, when the antibody can bind a multivalent antigen through more than one epitope simultaneously. A key goal of antibody engineering is thus to optimize avidity, but little is known about the nanoscale spatial dependence of avidity in antibodies. Here, we develop a set of anti-parallel coiled-coils spanning from 8-21 nm and validate their structure using biophysical techniques. We use the coiled-coils to control the spacing between two epitopes, and measure how antigen spacing affects the stability of the bivalent antibody:antigen complex. We find a maximal avidity enhancement at a spacing of 14 nm, but only see a ∼2-fold variation of avidity in the range from 8-21 nm. In contrast to recent studies, we find the avidity to be relatively insensitive to epitope spacing near the avidity maximum as long as it is within the spatial tolerance of the antibody. The coiled-coil systems developed here may prove a useful protein nanocaliper for profiling the spatial tolerance and avidity profile of bispecific antibodies.

scholarly journals Nanoscale spatial dependence of avidity in an IgG1 antibody

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Agnieszka Jendroszek ◽  
Magnus Kjaergaard
Keyword(s):  
Spatial Dependence ◽  
Target Recognition ◽  
Coiled Coil ◽  
Coiled Coils ◽  
Antibody Engineering ◽  
Bispecific Antibodies ◽  
Secreted Proteins ◽  
Biophysical Techniques ◽  
The Stability ◽  
20 Nm

AbstractAntibodies are secreted proteins that are crucial to recognition of pathogens by the immune system and are also efficient pharmaceuticals. The affinity and specificity of target recognition can increase remarkably through avidity effects, when the antibody can bind a multivalent antigen through more than one epitope simultaneously. A key goal of antibody engineering is thus to optimize avidity, but little is known about the nanoscale spatial dependence of avidity in antibodies. Here, we develop a set of anti-parallel coiled-coils spanning from 7 to 20 nm and validate their structure using biophysical techniques. We use the coiled-coils to control the spacing between two epitopes, and measure how antigen spacing affects the stability of the bivalent antibody:antigen complex. We find a maximal avidity enhancement at a spacing of 13 nm. In contrast to recent studies, we find the avidity to be relatively insensitive to epitope spacing near the avidity maximum as long as it is within the spatial tolerance of the antibody. We thus only see a ~ twofold variation of avidity in the range from 7 to 20 nm. The coiled-coil systems developed here may prove a useful protein nanocaliper for profiling the spatial tolerance and avidity profile of bispecific antibodies.

scholarly journals Coiled-Coil Trigger Motifs in the 1B and 2B Rod Domain Segments Are Required for the Stability of Keratin Intermediate Filaments

2000 ◽  
Vol 11 (10) ◽  
pp. 3539-3558 ◽  
Author(s):  
Kenneth C. Wu ◽  
Janine T. Bryan ◽  
Maria I. Morasso ◽  
Shyh-Ing Jang ◽  
Jeung-Hoon Lee ◽  
...  
Keyword(s):  
Coiled Coil ◽  
Coiled Coils ◽  
Detectable Effect ◽  
Keratin 5 ◽  
Molecular Stability ◽  
Potential Trigger ◽  
Helical Proteins ◽  
The Stability

Many α-helical proteins that form two-chain coiled coils possess a 13-residue trigger motif that seems to be required for the stability of the coiled coil. However, as currently defined, the motif is absent from intermediate filament (IF) protein chains, which nevertheless form segmented two-chain coiled coils. In the present work, we have searched for and identified two regions in IF chains that are essential for the stability necessary for the formation of coiled-coil molecules and thus may function as trigger motifs. We made a series of point substitutions with the keratin 5/keratin 14 IF system. Combinations of the wild-type and mutant chains were assembled in vitro and in vivo, and the stabilities of two-chain (one-molecule) and two-molecule assemblies were examined with use of a urea disassembly assay. Our new data document that there is a region located between residues 100 and 113 of the 2B rod domain segment that is absolutely required for molecular stability and IF assembly. This potential trigger motif differs slightly from the consensus in having an Asp residue at position 4 (instead of a Glu) and a Thr residue at position 9 (instead of a charged residue), but there is an absolute requirement for a Glu residue at position 6. Because these 13 residues are highly conserved, it seems possible that this motif functions in all IF chains. Likewise, by testing keratin IF with substitutions in both chains, we identified a second potential trigger motif between residues 79 and 91 of the 1B rod domain segment, which may also be conserved in all IF chains. However, we were unable to find a trigger motif in the 1A rod domain segment. In addition, many other point substitutions had little detectable effect on IF assembly, except for the conserved Lys-23 residue of the 2B rod domain segment. Cross-linking and modeling studies revealed that Lys-23 may lie very close to Glu-106 when two molecules are aligned in the A22 mode. Thus, the Glu-106 residue may have a dual role in IF structure: it may participate in trigger formation to afford special stability to the two-chain coiled-coil molecule, and it may participate in stabilization of the two-molecule hierarchical stage of IF structure.

scholarly journals Metabolic enzyme clustering by coiled coils improves the biosynthesis of resveratrol and mevalonate

2020 ◽  
Author(s):  
Tina Fink ◽  
Bojana Stevović ◽  
René Verwaal ◽  
Johannes A. Roubos ◽  
Rok Gaber ◽  
...  
Keyword(s):  
Spatial Organization ◽  
Noncovalent Interactions ◽  
Coiled Coil ◽  
Product Yield ◽  
Coiled Coils ◽  
Metabolic Enzyme ◽  
Biosynthetic Enzymes ◽  
Reaction Products ◽  
Interaction Domains ◽  
The Stability

Abstract The clustering of biosynthetic enzymes is used in nature to channel reaction products and increase the yield of compounds produced by multiple reaction steps. The coupling of multiple enzymes has been shown to increase the biosynthetic product yield. Different clustering strategies have particular advantages as the spatial organization of multiple enzymes creates biocatalytic cascades with a higher efficiency of biochemical reaction. However, there are also some drawbacks, such as misfolding and the variable stability of interaction domains, which may differ between particular biosynthetic reactions and the host organism. Here, we compared different protein-based clustering strategies, including direct fusion, fusion mediated by intein, and noncovalent interactions mediated through small coiled-coil dimer-forming domains. The clustering of enzymes through orthogonally designed coiled-coil interaction domains increased the production of resveratrol in Escherichia coli more than the intein-mediated fusion of biosynthetic enzymes. The improvement of resveratrol production correlated with the stability of the coiled-coil dimers. The coiled-coil fusion-based approach also increased mevalonate production in Saccharomyces cerevisiae , thus demonstrating the wider applicability of this strategy.

scholarly journals Metabolic enzyme clustering by coiled coils improves the biosynthesis of resveratrol and mevalonate

2020 ◽  
Author(s):  
Tina Fink ◽  
Bojana Stevović ◽  
René Verwaal ◽  
Johannes A. Roubos ◽  
Rok Gaber ◽  
...  
Keyword(s):  
Spatial Organization ◽  
Noncovalent Interactions ◽  
Coiled Coil ◽  
Product Yield ◽  
Coiled Coils ◽  
Metabolic Enzyme ◽  
Biosynthetic Enzymes ◽  
Reaction Products ◽  
Interaction Domains ◽  
The Stability

Abstract The clustering of biosynthetic enzymes is used in nature to channel reaction products and increase the yield of compounds produced by multiple reaction steps. The coupling of multiple enzymes has been shown to increase the biosynthetic product yield. Different clustering strategies have particular advantages as the spatial organization of multiple enzymes creates biocatalytic cascades with a higher efficiency of biochemical reaction. However, there are also some drawbacks, such as misfolding and the variable stability of interaction domains, which may differ between particular biosynthetic reactions and the host organism. Here, we compared different protein-based clustering strategies, including direct fusion, fusion mediated by intein, and noncovalent interactions mediated through small coiled-coil dimer-forming domains. The clustering of enzymes through orthogonally designed coiled-coil interaction domains increased the production of resveratrol in Escherichia coli more than the intein-mediated fusion of biosynthetic enzymes. The improvement of resveratrol production correlated with the stability of the coiled-coil dimers. The coiled-coil fusion-based approach also increased mevalonate production in Saccharomyces cerevisiae , thus demonstrating the wider applicability of this strategy.

scholarly journals Importance and Considerations of Antibody Engineering in Antibody-Drug Conjugates Development from a Clinical Pharmacologist’s Perspective

Antibodies ◽  
2021 ◽  
Vol 10 (3) ◽  
pp. 30
Author(s):  
Andrew T. Lucas ◽  
Amber Moody ◽  
Allison N. Schorzman ◽  
William C. Zamboni
Keyword(s):  
Immune System ◽  
Market Value ◽  
Therapeutic Index ◽  
Drug Carriers ◽  
Antibody Engineering ◽  
Antibody Drug Conjugates ◽  
Success Stories ◽  
Drug Conjugates ◽  
Early Success ◽  
Antibody Drug

Antibody-drug conjugates (ADCs) appear to be in a developmental boom, with five FDA approvals in the last two years and a projected market value of over $4 billion by 2024. Major advancements in the engineering of these novel cytotoxic drug carriers have provided a few early success stories. Although the use of these immunoconjugate agents are still in their infancy, valuable lessons in the engineering of these agents have been learned from both preclinical and clinical failures. It is essential to appreciate how the various mechanisms used to engineer changes in ADCs can alter the complex pharmacology of these agents and allow the ADCs to navigate the modern-day therapeutic challenges within oncology. This review provides a global overview of ADC characteristics which can be engineered to alter the interaction with the immune system, pharmacokinetic and pharmacodynamic profiles, and therapeutic index of ADCs. In addition, this review will highlight some of the engineering approaches being explored in the creation of the next generation of ADCs.

scholarly journals Coiled-coil networking shapes cell molecular machinery

2012 ◽  
Vol 23 (19) ◽  
pp. 3911-3922 ◽  
Author(s):  
Yongqiang Wang ◽  
Xinlei Zhang ◽  
Hong Zhang ◽  
Yi Lu ◽  
Haolong Huang ◽  
...  
Keyword(s):  
Protein Interactions ◽  
Critical Role ◽  
Coiled Coil ◽  
Coiled Coils ◽  
Protein Protein Interactions ◽  
Full Spectrum ◽  
Kinetochore Assembly ◽  
Genome Wide ◽  
Molecular Machinery ◽  
Systematic Understanding

The highly abundant α-helical coiled-coil motif not only mediates crucial protein–protein interactions in the cell but is also an attractive scaffold in synthetic biology and material science and a potential target for disease intervention. Therefore a systematic understanding of the coiled-coil interactions (CCIs) at the organismal level would help unravel the full spectrum of the biological function of this interaction motif and facilitate its application in therapeutics. We report the first identified genome-wide CCI network in Saccharomyces cerevisiae, which consists of 3495 pair-wise interactions among 598 predicted coiled-coil regions. Computational analysis revealed that the CCI network is specifically and functionally organized and extensively involved in the organization of cell machinery. We further show that CCIs play a critical role in the assembly of the kinetochore, and disruption of the CCI network leads to defects in kinetochore assembly and cell division. The CCI network identified in this study is a valuable resource for systematic characterization of coiled coils in the shaping and regulation of a host of cellular machineries and provides a basis for the utilization of coiled coils as domain-based probes for network perturbation and pharmacological applications.

scholarly journals ARCIMBOLDOon coiled coils

2018 ◽  
Vol 74 (3) ◽  
pp. 194-204 ◽  
Author(s):  
Iracema Caballero ◽  
Massimo Sammito ◽  
Claudia Millán ◽  
Andrey Lebedev ◽  
Nicolas Soler ◽  
...  
Keyword(s):  
Coiled Coil ◽  
Coiled Coils ◽  
Translational Symmetry ◽  
Command Line ◽  
Phase Problem ◽  
Final Solution ◽  
Resolution Limit ◽  
Helical Structures ◽  
Small Model ◽  
Test Pool

ARCIMBOLDOsolves the phase problem by combining the location of small model fragments usingPhaserwith density modification and autotracing usingSHELXE. Mainly helical structures constitute favourable cases, which can be solved using polyalanine helical fragments as search models. Nevertheless, the solution of coiled-coil structures is often complicated by their anisotropic diffraction and apparent translational noncrystallographic symmetry. Long, straight helices have internal translational symmetry and their alignment in preferential directions gives rise to systematic overlap of Patterson vectors. This situation has to be differentiated from the translational symmetry relating different monomers.ARCIMBOLDO_LITEhas been run on single workstations on a test pool of 150 coiled-coil structures with 15–635 amino acids per asymmetric unit and with diffraction data resolutions of between 0.9 and 3.0 Å. The results have been used to identify and address specific issues when solving this class of structures usingARCIMBOLDO. Features fromPhaserv.2.7 onwards are essential to correct anisotropy and produce translation solutions that will pass the packing filters. As the resolution becomes worse than 2.3 Å, the helix direction may be reversed in the placed fragments. Differentiation between true solutions and pseudo-solutions, in which helix fragments were correctly positioned but in a reverse orientation, was found to be problematic at resolutions worse than 2.3 Å. Therefore, after every new fragment-placement round, complete or sparse combinations of helices in alternative directions are generated and evaluated. The final solution is once again probed by helix reversal, refinement and extension. To conclude, density modification andSHELXEautotracing incorporating helical constraints is also exploited to extend the resolution limit in the case of coiled coils and to enhance the identification of correct solutions. This study resulted in a specialized mode withinARCIMBOLDOfor the solution of coiled-coil structures, which overrides the resolution limit and can be invoked from the command line (keyword coiled_coil) orARCIMBOLDO_LITEtask interface inCCP4i.

Cell-Penetrating Peptide-Mediated Nanovaccine Delivery

2021 ◽  
Vol 22 ◽  
Author(s):  
Jizong Jiang
Keyword(s):  
Immune System ◽  
Immune Responses ◽  
Cell Penetrating Peptide ◽  
Antigen Delivery ◽  
Efficient Manner ◽  
Antigen Uptake ◽  
Cell Penetrating ◽  
The Stability ◽  
Antigen Presenting

Abstract: Vaccination with small antigens, such as proteins, peptides, or nucleic acids, is used to activate the immune system and trigger the protective immune responses against a pathogen. Currently, nanovaccines are undergoing development instead of conventional vaccines. The size of nanovaccines is in the range of 10–500 nm, which enables them to be readily taken up by cells and exhibit improved safety profiles. However, low-level immune responses, as the removal of redundant pathogens, trigger counter-effective activation of the immune system invalidly and present a challenging obstacle to antigen recognition and its uptake via antigen-presenting cells (APCs). In addition, toxicity can be substantial. To overcome these problems, a variety of cell-penetrating peptide (CPP)-mediated vaccine delivery systems based on nanotechnology have been proposed, most of which are designed to improve the stability of antigens in vivo and their delivery into immune cells. CPPs are particularly attractive components of antigen delivery. Thus, the unique translocation property of CPPs ensures that they remain an attractive carrier with the capacity to deliver cargo in an efficient manner for the application of drugs, gene transfer, protein, and DNA/RNA vaccination delivery. CPP-mediated nanovaccines can enhance antigen uptake, processing, and presentation by APCs, which are the fundamental steps in initiating an immune response. This review describes the different types of CPP-based nanovaccines delivery strategies.

Hopf Bifurcation Induced by Delay Effect in a Diffusive Tumor-Immune System

2018 ◽  
Vol 28 (11) ◽  
pp. 1850136 ◽  
Author(s):  
Ben Niu ◽  
Yuxiao Guo ◽  
Yanfei Du
Keyword(s):  
Immune System ◽  
Hopf Bifurcation ◽  
Periodic Solutions ◽  
Center Manifold ◽  
Immune Cell ◽  
Tumor Treatment ◽  
Delay Effect ◽  
Stable Periodic ◽  
The Stability ◽  
Bifurcation And Stability

Tumor-immune interaction plays an important role in the tumor treatment. We analyze the stability of steady states in a diffusive tumor-immune model with response and proliferation delay [Formula: see text] of immune system where the immune cell has a probability [Formula: see text] in killing tumor cells. We find increasing time delay [Formula: see text] destabilizes the positive steady state and induces Hopf bifurcations. The criticality of Hopf bifurcation is investigated by deriving normal forms on the center manifold, then the direction of bifurcation and stability of bifurcating periodic solutions are determined. Using a group of parameters to simulate the system, stable periodic solutions are found near the Hopf bifurcation. The effect of killing probability [Formula: see text] on Hopf bifurcation values is also discussed.

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