Camelid and shark single domain antibodies: structural features and therapeutic potential

Könning, Doreen; Zielonka, Stefan; Grzeschik, Julius; Empting, Martin; Valldorf, Bernhard; Krah, Simon; Schröter, Christian; Sellmann, Carolin; Hock, Björn; Kolmar, Harald

doi:10.1016/j.sbi.2016.10.019

Cited by 167 publications

(131 citation statements)

References 63 publications

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“…Our results further demonstrate that by trapping specific states, nanobodies are useful for structural and mechanistic studies of ABC transporters. Investigation of these types of inhibitory molecules is a promising area of biology, which may lead to design innovative strategies for the treatment of human affections313233.…”

Section: Discussionmentioning

confidence: 99%

Structural basis of inhibition of lipid-linked oligosaccharide flippase PglK by a conformational nanobody

Pérez

Köhler

Janser

et al. 2017

Sci Rep

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PglK is an ABC transporter that flips a lipid-linked oligosaccharide (LLO) that serves as a donor in protein N-glycosylation. Previous structures revealed two inward-facing conformations, both with very large separations of the nucleotide binding domains (NBDs), and a closed, ADP-bound state that featured an occluded cavity. To investigate additional states, we developed conformation-sensitive, single-domain camelid nanobodies (Nb) and studied their effect on PglK activity. Biochemical, structural, and mass spectrometric analyses revealed that one inhibitory Nb binds as a single copy to homodimeric PglK. The co-crystal structure of this Nb and ADP-bound PglK revealed a new, narrowly inward-open conformation. Rather than inducing asymmetry in the PglK homodimer, the binding of one Nb results in steric constraints that prevent a second Nb to access the symmetry-related site in PglK. The Nb performed its inhibitory role by a “sticky-doorstop” mechanism, where inhibition of ATP hydrolysis and LLO flipping activity occurs due to impaired closing of the NBD interface, which prevents PglK from converting to an outward-open conformation. This inhibitory mode suggests tight conformational coupling between the ATPase sites, which may apply to other ABC transporters.

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Section: Discussionmentioning

confidence: 99%

Structural basis of inhibition of lipid-linked oligosaccharide flippase PglK by a conformational nanobody

Pérez

Köhler

Janser

et al. 2017

Sci Rep

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“…[2,3] Various smaller formats, including single chain variable fragments (scFv), antigen binding fragments (Fab), minibodies etc. [4] as well as antibody structures from alternative sources like camelid antibodies or immunoglobulin new antigen receptors [5] were investigated for their potential use as therapeutics. Additional efforts at engineering were made to create bispecific types of structures that can target two different antigens, [6] or even more recently, to produce antibodies that contain multiple crystallizable fragment (Fc)-domains, showing enhanced effector functions.…”

Section: Microheterogeneitymentioning

confidence: 99%

Microheterogeneity of Recombinant Antibodies: Analytics and Functional Impact

Beyer

Schuster

Jungbauer

et al. 2017

Biotechnology Journal

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Antibodies are typical examples of biopharmaceuticals which are composed of numerous, almost infinite numbers of potential molecular entities called variants or isoforms, which constitute the microheterogeneity of these molecules. These variants are generated during biosynthesis by so‐called posttranslational modification, during purification or upon storage. The variants differ in biological properties such as pharmacodynamic properties, for example, Antibody Dependent Cellular Cytotoxicity, complement activation, and pharmacokinetic properties, for example, serum half‐life and safety. Recent progress in analytical technologies such as various modes of liquid chromatography and mass spectrometry has helped to elucidate the structure of a lot of these variants and their biological properties. In this review the most important modifications (glycosylation, terminal modifications, amino acid side chain modifications, glycation, disulfide bond variants and aggregation) are reviewed and an attempt is made to give an overview on the biological properties, for which the reports are often contradictory. Even though there is a deep understanding of cellular and molecular mechanism of antibody modification and their consequences, the clinical proof of the effects observed in vitro and in vivo is still not fully rendered. For some modifications such as core‐fucosylation of the N‐glycan and aggregation the effects are clear and should be monitored, but with others such as C‐terminal lysine clipping the reports are contradictory. As a consequence it seems too early to tell if any modification can be safely ignored.

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“…If the antibody is obtained from camelids, a unique antibody protein known as a single-chain antibody or nanobody can be obtained [34]. A typical antibody usually consists of four polypeptide chains per molecule, whereas a camelid antibody has two chains per antibody molecule.…”

Section: Purification and Analysismentioning

confidence: 99%

A Simple Outline of Methods for Protein Isolation and Purification

Lee

2017

Endocrinol Metab

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At the summer workshop of the Korean Endocrine Society held in 2016, some examples of protein experiments were discussed in the session entitled “All about working with proteins.” In contrast to what the title suggested, it was unrealistic to comprehensively discuss all protein analytical methods. Therefore, the goal was to outline protein experimental techniques that are useful in research or in bench work. In conversations with clinicians, however, I have always felt that researchers who do not engage in bench science have different demands than those who do. Protein research tools that are useful in bench science may not be very useful or effective in the diagnostic field. In this paper, I provide a general summary of the protein analytical methods that are used in basic scientific research, and describe how they can be applied in the diagnostic field.

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Camelid and shark single domain antibodies: structural features and therapeutic potential

Cited by 167 publications

References 63 publications

Structural basis of inhibition of lipid-linked oligosaccharide flippase PglK by a conformational nanobody

Structural basis of inhibition of lipid-linked oligosaccharide flippase PglK by a conformational nanobody

Microheterogeneity of Recombinant Antibodies: Analytics and Functional Impact

A Simple Outline of Methods for Protein Isolation and Purification

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