Alternative Protein Scaffolds as Novel Biotherapeutics

Gebauer, Michaela; Skerra, Arne

doi:10.1007/978-1-4939-2543-8_13

Cited by 9 publications

(11 citation statements)

References 242 publications

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“…Based on repeat protein domains, these proteins are simple to design and modular, with highly variable binding surfaces. Compared with immunoglobulins, these proteins offer high affinity and specificity for their targets . While many protein engineering efforts are only able to target proteins and peptides, these engineered immune proteins are able to target glycans as well, filling an important gap in tools for research and biomedical applications.…”

Section: Discussionmentioning

confidence: 99%

“…Further, they are comparatively easy to design, have highly variable binding surfaces, and can be remarkably stable . Engineered binding scaffolds based on repeat proteins have found applications in biosensing, diagnostics, and therapeutics …”

Section: Engineering Repeat Proteinsmentioning

confidence: 99%

“…Immunoglobulin (Ig) antibodies are the best‐studied examples of such recognition agents, and have been reviewed extensively . However, problems with stability, expense of expression, and specificity of interaction of Ig antibodies have led in recent years to the development of alternative binding scaffolds . Antibodies rely on disulfide bond formation for their stability, requiring difficult, expensive eukaryotic cell production; this also makes them unsuitable for applications in reducing conditions .…”

Section: Introductionmentioning

confidence: 99%

“…In the search for better recognition agents, many different scaffolds have emerged; some notable examples are peptide and DNA aptamers, affimers, and anticalins, among others. The motivation and results of this research have been extensively reviewed in recent years . among these scaffolds, repeat proteins such as designed ankyrin repeat proteins (DARPins) tetratricopeptide repeat proteins (TPRs), and armadillo repeats have emerged as a promising class of scaffolds.…”

Section: Introductionmentioning

confidence: 99%

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Engineering repeat proteins of the immune system

McCord

Grove

2020

Biopolymers

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Limitations associated with immunoglobulins have motivated the search for novel binding scaffolds. Repeat proteins have emerged as one promising class of scaffolds, but often are limited to binding protein and peptide targets. An exception is the repeat proteins of the immune system, which have in recent years served as an inspiration for binding scaffolds which can bind glycans and other classes of biomolecule.Like other repeat proteins, these proteins can be very stable and have a monomeric mode of binding, with elongated and highly variable binding surfaces. The ability to target glycans and glycoproteins fill an important gap in current tools for research and biomedical applications.

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Section: Discussionmentioning

confidence: 99%

Section: Engineering Repeat Proteinsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Engineering repeat proteins of the immune system

McCord

Grove

2020

Biopolymers

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“…More than 50 [15] APSs have been proposed to date [1–9, 13], numerous ABPs are in clinical trials for treatment of neoplastic, autoimmune, inflammatory, infectious, and ophthalmological diseases [8, 9, 16–19], and one ABP, ecallantide (KALBITOR ® ), has already reached pharmaceutical market. Although several APSs (such as 10 th human fibronectin type III domain, Fc-binding Z domain derived from staphylococcal protein A, lipocalins and ankyrin fold) are already broadly established APSs, the natural process of evolution of artificial binding proteins will witness extension of their applications, polishing of validated APSs and development of novel protein scaffolds with superior properties.…”

Section: Introductionmentioning

confidence: 99%

Derivative of Extremophilic 50S Ribosomal Protein L35Ae as an Alternative Protein Scaffold

Lomonosova

Улитин²,

Kazakov

et al. 2017

PLoS ONE

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Small antibody mimetics, or alternative binding proteins (ABPs), extend and complement antibody functionality with numerous applications in research, diagnostics and therapeutics. Given the superiority of ABPs, the last two decades have witnessed development of dozens of alternative protein scaffolds (APSs) for the design of ABPs. Proteins from extremophiles with their high structural stability are especially favorable for APS design. Here, a 10X mutant of the 50S ribosomal protein L35Ae from hyperthermophilic archaea Pyrococcus horikoshii has been probed as an APS. A phage display library of L35Ae 10X was generated by randomization of its three CDR-like loop regions (repertoire size of 2×108). Two L35Ae 10X variants specific to a model target, the hen egg-white lysozyme (HEL), were isolated from the resulting library using phage display. The affinity of these variants (L4 and L7) to HEL ranges from 0.10 μM to 1.6 μM, according to surface plasmon resonance data. While L4 has 1–2 orders of magnitude lower affinity to HEL homologue, bovine α-lactalbumin (BLA), L7 is equally specific to HEL and BLA. The reference L35Ae 10X is non-specific to both HEL and BLA. L4 and L7 are more resistant to denaturation by guanidine hydrochloride compared to the reference L35Ae 10X (mid-transition concentration is higher by 0.1–0.5 M). Chemical crosslinking experiments reveal an increased propensity of L4 and L7 to multimerization. Overall, the CDR-like loop regions of L35Ae 10X represent a proper interface for generation of functional ABPs. Hence, L35Ae is shown to extend the growing family of protein scaffolds dedicated to the design of novel binding proteins.

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Fusion with an albumin‐binding domain improves pharmacokinetics of an αvβ3‐integrin binding fibronectin scaffold protein

Gapizov

Petrovskaya

Shingarova

et al. 2019

Biotech and App Biochem

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Fusion with an albumin-binding domain (ABD) of streptococcal protein G represents a popular approach for half-life extension of small protein therapeutics in the organism. To increase the circulation time of engineered αvβ3-integrin-binding protein (JCL) based on the 10th human fibronectin type III domain ( 10 Fn3), we have constructed several fusions with ABD with different orientations of the partner proteins and linker length. The recombinant proteins were expressed in Escherichia coli cells and purified by nickel-affinity chromatography. All fusion proteins bound human serum albumin (HSA) in ELISA assay; however, fusions with longer linkers demonstrated better performance. Interaction of ABD-L 15 -JCL and JCL-L 14 -ABD with HSA was confirmed by analytical size exclusion chromatography and pull-down assays. Surprisingly, the thermal stability of ABD-L 15 -JCL was dramatically decreased in comparison with JCL and JCL-L 14 -ABD proteins. Pharmacokinetic studies revealed that JCL-L 14 -ABD circulated in murine blood about 10 times longer than ABD-L 15 -JCL and 960 times longer than JCL. Biodistribution studies of JCL-L 14 -ABD in mice revealed its increased level in blood and a decreased accumulation in liver and kidneys in comparison with JCL. Obtained results demonstrate the utility of the fusion with ABD for half-life extension of the binding proteins based on 10

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Alternative Protein Scaffolds as Novel Biotherapeutics

Cited by 9 publications

References 242 publications

Engineering repeat proteins of the immune system

Engineering repeat proteins of the immune system

Derivative of Extremophilic 50S Ribosomal Protein L35Ae as an Alternative Protein Scaffold

Fusion with an albumin‐binding domain improves pharmacokinetics of an αvβ3‐integrin binding fibronectin scaffold protein

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