Of all the antibody classes, IgE displays a uniquely slow dissociation rate from, and high affinity for, its cell surface receptor FcεRI. The structural basis for these key determinants of IgE's ability to mediate allergic hypersensitivity is now revealed by the 3.4Å resolution crystal structure of human IgE-Fc (consisting of the Cε2, Cε3 and Cε4 domains) bound to the extracellular domains of the FcεRI α-chain. Comparison with free IgE-Fc (reported here at 1.9Å) shows that the antibody, which has a compact, bent structure prior to receptor engagement, becomes even more acutely bent in the complex. Thermodynamic analysis indicates that the interaction is entropically driven, which explains how the non-contacting Cε2 domains, in place of the flexible hinge region of IgG antibodies, contribute together with the conformational changes to IgE's unique binding properties.The global incidence of allergic disease has increased markedly in recent years and continues to rise. Asthma currently affects 22.2 million people in North America and 5.4 million in the UK where the rate of incidence, especially in children, is among the highest in the world 1 . Anaphylactic reactions to foods such as nuts, virtually unknown thirty years ago, are now relatively common. The reason for this increase is debated, but these and other allergic conditions (rhinitis, eczema etc.) are all mediated by IgE, and the viability of AUTHOR CONTRIBUTIONS MDH and AMD carried out the crystallographic analysis of the complex, and BD the crystallographic analysis of IgE-Fc; MDH, JEN, JH, AJB and RJO produced the proteins; SCB and JMM carried out the thermodynamic analysis; EG contributed to the analysis of the conformational changes; HJG, AJB and BJS planned and directed the project; MDH, AMD, JMM and BJS wrote the paper.Accession codes. Atomic coordinates and structure factors have been deposited in the Protein Data Bank with entry codes 2WQR for IgE-Fc and 2Y7Q for the complex. [6][7][8][9] . Particular interest has centered on the role of the Cε2 domains, which were shown to contribute to the slow dissociation rate 10 , by comparing the binding kinetics of the complete IgE-Fc (dimer Cε2-Cε3-Cε4 domains) with a sub-fragment lacking the Cε2 domains, here referred to as Fcε3-4. The Cε2 domains have no counterpart in IgG, all four subclasses of which have in their place hinge regions of various lengths and degrees of flexibility. Europe PMC Funders GroupEarly fluorescence resonance energy transfer (FRET) studies of labelled chimeric IgE indicated a more compact and bent structure than the extended, flexible Y-shaped IgG 11 , and later solution scattering studies of IgE-Fc were consistent with such a structure 12 . The crystal structure of IgE-Fc revealed for the first time the extent and nature of this bend 13 , made possible by the presence of the Cε2 domains. Surprisingly the molecule was found to be so acutely and asymmetrically bent, with the Cε2 domain pair folded back across the Cε3 domains, that one Cε2 domain even contacted the Cε4 domai...

Human IgG4, normally the least abundant of the four subclasses of IgG in serum, displays a number of unique biological properties. It can undergo heavy-chain exchange, also known as Fab-arm exchange, leading to the formation of monovalent but bispecific antibodies, and it interacts poorly with FcγRII and FcγRIII, and complement. These properties render IgG4 relatively “non-inflammatory” and have made it a suitable format for therapeutic monoclonal antibody production. However, IgG4 is also known to undergo Fc-mediated aggregation and has been implicated in auto-immune disease pathology. We report here the high-resolution crystal structures, at 1.9 and 2.35 Å, respectively, of human recombinant and serum-derived IgG4-Fc. These structures reveal conformational variability at the CH3–CH3 interface that may promote Fab-arm exchange, and a unique conformation for the FG loop in the CH2 domain that would explain the poor FcγRII, FcγRIII and C1q binding properties of IgG4 compared with IgG1 and -3. In contrast to other IgG subclasses, this unique conformation folds the FG loop away from the CH2 domain, precluding any interaction with the lower hinge region, which may further facilitate Fab-arm exchange by destabilisation of the hinge. The crystals of IgG4-Fc also display Fc–Fc packing contacts with very extensive interaction surfaces, involving both a consensus binding site in IgG-Fc at the CH2–CH3 interface and known hydrophobic aggregation motifs. These Fc–Fc interactions are compatible with intact IgG4 molecules and may provide a model for the formation of aggregates of IgG4 that can cause disease pathology in the absence of antigen.

The crystal structures of thymidine kinase from herpes simplex virus type-1 complexed with its natural substrate deoxythymidine (dT) and complexed with the guanosine analogue Ganciclovir have been solved. Both structures are in the C222(1) crystal form with two molecules per asymmetric unit related by a non-crystallographic two-fold axis. The present models have been refined to 2.8 A and 2.2 A, with crystallographic R factors of 24.1% and 23.3% for the dT and Ganciclovir complexes respectively, without the inclusion of any solvent molecules. The core of the molecule exhibits high structural homology with adenylate kinase and other nucleotide binding proteins. These structural similarities provide an insight into the mechanism of nucleoside phosphorylation by thymidine kinase.

IgG4, the least represented human IgG subclass in serum, is an intriguing antibody with unique biological properties, such as the ability to undergo Fab-arm exchange and limit immune complex formation. The lack of effector functions, such as antibody-dependent cell-mediated cytotoxicity and complement-dependent cytotoxicity, is desirable for therapeutic purposes. IgG4 plays a protective role in allergy by acting as a blocking antibody, and inhibiting mast cell degranulation, but a deleterious role in malignant melanoma, by impeding IgG1-mediated anti-tumor immunity. These findings highlight the importance of understanding the interaction between IgG4 and Fcγ receptors. Despite a wealth of structural information for the IgG1 subclass, including complexes with Fcγ receptors, and structures for intact antibodies, high-resolution crystal structures were not reported for IgG4-Fc until recently. Here, we highlight some of the biological properties of human IgG4, and review the recent crystal structures of IgG4-Fc. We discuss the unexpected conformations adopted by functionally important Cγ2 domain loops, and speculate about potential implications for the interaction between IgG4 and FcγRs.

IgG4 is the least abundant subclass of IgG in normal human serum, but elevated IgG4 levels are triggered in response to a chronic antigenic stimulus and inflammation. Since the immune system is exposed to tumor-associated antigens over a relatively long period of time, and tumors notoriously promote inflammation, it is unsurprising that IgG4 has been implicated in certain tumor types. Despite differing from other IgG subclasses by only a few amino acids, IgG4 possesses unique structural characteristics that may be responsible for its poor effector function potency and immunomodulatory properties. We describe the unique attributes of IgG4 that may be responsible for these regulatory functions, particularly in the cancer context. We discuss the inflammatory conditions in tumors that support IgG4, the emerging and proposed mechanisms by which IgG4 may contribute to tumor-associated escape from immune surveillance and implications for cancer immunotherapy.

Immunoglobulin E (IgE) is well known for its role in allergic disease, the manifestations of which are mediated through its two Fc receptors, FcεRI and CD23 (FcεRII). IgE and its interactions with these receptors are therefore potential targets for therapeutic intervention, and exciting progress has been made in this direction. Furthermore, recent structural studies of IgE-Fc, the two receptors, and of their complexes, have revealed a remarkable degree of plasticity at the IgE-CD23 interface and an even more remarkable degree of dynamic flexibility within the IgE molecule. Indeed, there is allosteric communication between the two receptor-binding sites, which we now know are located at some distance from each other in IgE-Fc (at opposite ends of the Cε3 domain). The conformational changes associated with FcεRI and CD23 binding to IgE-Fc ensure that their interactions are mutually incompatible, and it may be that this functional imperative has driven IgE to evolve such a dynamic structure. Appreciation of these new structural data has revised our view of IgE structure, shed light on the co-evolution of antibodies and their receptors, and may open up new therapeutic opportunities.

Background: Fab arm exchange requires weak interactions between CH3 domains, such as in human IgG4.Results: CH3-CH3 interactions differ >1,000,000-fold between human subclasses and allotypes due to variations Lys/Asn-392, Val/Met-397, and Lys/Arg-409.Conclusion: For IgG2 and IgG3, but not IgG1, hinge disulfide bonds are essential to prevent half-molecule dissociation.Significance: Subclass/allotype variation in the CH3 domain can alter antibody stability and functionality.

Immunoglobulin E and its interactions with receptors FcϵRI and CD23 play a central role in allergic disease. Omalizumab, a clinically approved therapeutic antibody, inhibits the interaction between IgE and FcϵRI, preventing mast cell and basophil activation, and blocks IgE binding to CD23 on B cells and antigen-presenting cells. We solved the crystal structure of the complex between an omalizumab-derived Fab and IgE-Fc, with one Fab bound to each Cϵ3 domain. Free IgE-Fc adopts an acutely bent structure, but in the complex it is only partially bent, with large-scale conformational changes in the Cϵ3 domains that inhibit the interaction with FcϵRI. CD23 binding is inhibited sterically due to overlapping binding sites on each Cϵ3 domain. Studies of omalizumab Fab binding in solution demonstrate the allosteric basis for FcϵRI inhibition and, together with the structure, reveal how omalizumab may accelerate dissociation of receptor-bound IgE from FcϵRI, exploiting the intrinsic flexibility and allosteric potential of IgE.