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Human immunoglobulin G (IgG) exists as four subclasses IgG1-4, each of which has two Fab subunits joined by two hinges to a Fc subunit. IgG4 has the shortest hinge with 12 residues. The Fc subunit has two glycan chains, but the importance of glycosylation is not fully understood in IgG4. Here, to evaluate the stability and structure of non-glycosylated IgG4, we performed a multidisciplinary structural study of glycosylated and deglycosylated human IgG4 A33 for comparison with our similar study of human IgG1 A33. After deglycosylation, IgG4 was found to be monomeric by analytical ultracentrifugation; its sedimentation coefficient of 6.52 S was reduced by 0.27 S in reflection of its lower mass. X-ray and neutron solution scattering showed that the overall Guinier radius of gyration RG and its cross-sectional values after deglycosylation were almost unchanged. In the P(r) distance distribution curves, the two M1 and M2 peaks that monitor the two most common distances within IgG4 were unchanged following deglycosylation. Further insight from Monte Carlo simulations for glycosylated and deglycosylated IgG4 came from 111,382 and 117,135 possible structures respectively. Their comparison to the X-ray and neutron scattering curves identified several hundred best-fit models for both forms of IgG4. Principal component analyses showed that glycosylated and deglycosylated IgG4 exhibited different conformations from each other. Within the constraint of unchanged RG and M1-M2 values, the glycosylated IgG4 models showed more restricted Fc conformations compared to deglycosylated IgG4, but no other changes. Kratky plots supported this interpretation of greater disorder upon deglycosylation, also observed in IgG1. Overall, these more variable Fc conformations may demonstrate a generalisable impact of deglycosylation on Fc structures, but with no large conformational changes in IgG4 unlike those seen in IgG1. [ABSTRACT FROM AUTHOR]

Purpose: The purpose of this paper is to examine how and why individuals involved in executive remuneration (top pay) decision-making consider quantum as being appropriate rather than excessive, theorised under the rubric of accountability. Design/methodology/approach: In-depth interviews were conducted with non-executive directors (NEDs) serving on remuneration committees (Remcos), institutional investors, their external advisers and internal HR reward experts. Transcripts were analysed using NVivo and the Gioia qualitative methodology. Findings: Defining, measuring and applying performance conditionality in the determination of top pay quantum such that it aligns with company strategy/culture and values, as well as individual recipient motivations, is difficult. While creative approaches to setting top pay so as to attract, retain and motivate key personnel are welcomed, these risk Remco members' personal/organisational reputations. Members recognise disconnection between top pay quantum and general pay levels and how the media highlights social inequality leading to public distrust. They believe they can contribute to more socially acceptable quantum by applying their own values in top pay decision-making. Originality/value: Sanctions-based, trust-based and selection/peer networks/felt-based accountability theory is used to explain decision-makers' actions when determining top pay quantum. This paper extends felt accountability theory to encompass public/societal accountability in the context of the appropriateness of top pay quantum decisions. [ABSTRACT FROM AUTHOR]

FcγRI (CD64) is the only high-affinity Fcγ receptor found on monocytes, macrophages, eosinophils, neutrophils and dendritic cells. It binds immunoglobulin G (IgG) antibody-antigen complexes at its Fc region to trigger key immune responses. CD64 contains three immunoglobulin-fold extracellular domains (D1, D2 and D3) and a membrane-spanning region. Despite the importance of CD64, no solution structure for this is known to date. To investigate this, we used analytical ultracentrifugation, small-angle X-ray scattering, and atomistic modelling. Analytical ultracentrifugation revealed that CD64 was monomeric with a sedimentation coefficient s020,w of 2.53 S, together with some dimer. Small-angle X-ray scattering showed that its radius of gyration RG was 3.3–3.4 nm and increased at higher concentrations to indicate low dimerization. Monte Carlo modelling implemented in the SASSIE-web package generated 279,162 physically-realistic trial CD64 structures. From these, the scattering best-fit models at the lowest measured concentrations that minimised dimers revealed that the D1, D2 and D3 domains were structurally similar to those seen in three CD64 crystal structures, but showed previously unreported flexibility between D1, D2 and D3. Despite the limitations of the scattering data, the superimposition of the CD64 solution structures onto crystal structures of the IgG Fc-CD64 complex showed that the CD64 domains do not sterically clash with the IgG Fc region, i.e. the solution structure of CD64 was sufficiently compact to allow IgG to bind to its high-affinity Fcγ receptor. This improved understanding may result in novel approaches to inhibit CD64 function, and opens the way for the solution study of the full-length CD64-IgG complex. [ABSTRACT FROM AUTHOR]

Purpose--The purpose of this article is to enhance understanding of influences on interaction between corporate personnel and development specialists and line functions associated with expatriating managers. Line managers are expected to accept greater responsibility for people management and development. But line managers' strategies for managing risks inherent in supervising expatriate managers may cause to surface incompatibilities with specialists' corporate "policy conscience" role. A pluralistically inclined perspective on "managerial interest streams" offers insights into inter-group perceptions and behaviour. Design/methodology/approach--Focusing on organisational actors' interpretations, a non-standardised survey by e-mail, covering a small sample of expatriate managers (n=20) employed in various countries by a large UK-headquartered healthcare retail group, was complemented by semi-structured interviews with personnel and development specialists in a further seven large multinational companies. Findings--Potential tensions around the application of corporate expatriation policy may be attributed to factors "educating" line and specialist orientations to expatriate managers. Originality/value--The value of the paper is in the development of an original model sketching pluralistically located interaction around expatriation management. While limited to an exploratory empirical investigation, the practical implications derive from specification of opportunities and threats to partnership building between those involved in expatriating managers. (Contains 1 figure.)

ZAG is a multifunctional glycoprotein with a class I MHC‐like protein fold and an α1‐α2 lipid‐binding groove. The intrinsic ZAG ligand is unknown. Our previous studies showed that ZAG binds the dansylated C11 fatty acid, DAUDA, differently to the boron dipyrromethane C16 fatty acid, C16‐BODIPY. Here, the molecular basis for this difference was elucidated. Multi‐wavelength analytical ultracentrifugation confirmed that DAUDA and C16‐BODIPY individually bind to ZAG and compete for the same binding site. Molecular docking of lipid‐binding in the structurally related Cluster of differentiation 1 proteins predicted nine conserved ligand contact residues in ZAG. Twelve mutants were accordingly created by alanine scanning site directed mutagenesis for characterisation. Mutation of Y12 caused ZAG to misfold. Mutation of K147, R157 and A158 abrogated C16‐BODIPY but not DAUDA binding. L69 and T169 increased the fluorescence emission intensity of C16‐BODIPY but not of DAUDA compared to wild‐type ZAG and showed that C16‐BODIPY binds close to T169 and L69. Distance measurements of the crystal structure revealed K147 forms a salt bridge with D83. A range of bioactive bulky lipids including phospholipids and sphingolipids displaced DAUDA from the ZAG binding site but unexpectedly did not displace C16‐BODIPY. We conclude that the ZAG α1‐α2 groove contains separate but overlapping sites for DAUDA and C16‐BODIPY and is involved in binding to a bulkier and wider repertoire of lipids than previously reported. This work suggested that the in vivo activity of ZAG may be dictated by its lipid ligand. [ABSTRACT FROM AUTHOR]

Results demonstrate that the short consensus/complement repeat 1-2 domains are involved in the primary C3dg ligand binding site of human complement receptor CR2 with additional short domains of CR2 facilitating interaction with the ligand. Data reveal that His residue between the two short domains may be influencing the complex formation.

The human complement factor I is a plasma serine protease that contains five domains consisting of One factor I-membrane attack complex domain, one CD5 domain, two low-density lipoprotein receptor domains and one serine protease domain. Factor one is essential in the regulation of complement. X-ray and neutron scattering were undertaken to determine the arrangement of these domain on complement factor I derived from human plasma.

The structure of the heterodimeric receptor-ligand complex found in human coagulation factor VIIa (FVIIa) has been studied using x-ray and neutron scattering. FVIIa is a soluble four-domain plasma serine protease that binds with the two extracellular domains of the transmembrane tissue factor during blood coagulation. The study gives insight on the domain structure of free FVIIa in solution to illustrate how membrane-bound tissue factors detect the FVIIa as a ligand.

X-ray and neutron scattering methods were utilized to determine the solution structure of AmiC and the occurrence of a ligand-dependent conformational change. Results have shown that AmiC behaved differently from the typical periplasmic proteins such that it formed antiparallel dimers in its crystal structure secondary to the crystal lattice formation. Furthermore, it was shown that amiC participated in a monomer-trimer association in the solution.

There are fourteen domains in human immunoglobulin E (IgE) having specific fold structures. IgE contains two domains more than IgG in the Fc region. This makes the number of domains in the IgE-Fc six against four in the IgG-Fc. The bent models of IgE-Fc, with the additional Fc domain pair rotating by 40-50 degrees, produce the best agreement with X-ray and neutron scattering curves. The bent model is also in line with the hydrodynamic model of IgE.

Analysis of the interaction between immunoglobulin E (IgE) and its high-affinity receptor Fc-epsilon-RI reveals the formation of an equimolar complex between IgE-Fc and a soluble fragment of the Fc-epsilon-RI-alpha chain. Sedimentation equilibrium-derived molecular weight of the complex confirms the 1:1 stoichiometry, excluding all highly extended geometries for the complex.

The overall structure of C9 of human complement was determined by neutron and X-ray solution scattering techniques. It was shown that the solution structure of C9 is more elongated than previously deduced from electron microscopy studies and has a maximum dimension estimated as 12 +/- 2 nm. The scattering data were compatible with a limited range of arrangements of the three domains and perforin-like region in monomeric C9. Moreover, the structure modeling for C9 suggests that these four regions are arranged in a V-shaped structure, with an angle of 10 degrees between the two arms, each of length 11.1 nm.

Complement Factor H (CFH), with 20 short complement regulator (SCR) domains, regulates the alternative pathway of complement in part through the interaction of its C-terminal SCR-19 and SCR-20 domains with host cell-bound C3b and anionic oligosaccharides. In solution, CFH forms small amounts of oligomers, with one of its self-association sites being in the SCR-16/20 domains. In order to correlate CFH function with dimer formation and the occurrence of rare disease-associated variants in SCR-16/20, we identified the dimerization site in SCR-16/20. For this, we expressed, in Pichia pastoris , the five domains in SCR-16/20 and six fragments of this with one-three domains (SCR-19/20, SCR-18/20, SCR-17/18, SCR-16/18, SCR-17 and SCR-18). Size-exclusion chromatography suggested that SCR dimer formation occurred in several fragments. Dimer formation was clarified using analytical ultracentrifugation, where quantitative c(s) size distribution analyses showed that SCR-19/20 was monomeric, SCR-18/20 was slightly dimeric, SCR-16/20, SCR-16/18 and SCR-18 showed more dimer formation, and SCR-17 and SCR-17/18 were primarily dimeric with dissociation constants of ~5 µM. The combination of these results located the SCR-16/20 dimerization site at SCR-17 and SCR-18. X-ray solution scattering experiments and molecular modelling fits confirmed the dimer site to be at SCR-17/18, this dimer being a side-by-side association of the two domains. We propose that the self-association of CFH at SCR-17/18 enables higher concentrations of CFH to be achieved when SCR-19/20 are bound to host cell surfaces in order to protect these better during inflammation. Dimer formation at SCR-17/18 clarified the association of genetic variants throughout SCR-16/20 with renal disease. [ABSTRACT FROM AUTHOR]

Hereditary blood coagulation factor VII (FVII) deficiency is a rare autosomal recessive bleeding disorder resulting from variants in the gene encoding FVII (F7). Integration of genetic variation with functional consequences on protein function is essential for the interpretation of the pathogenicity of novel variants. Here, we describe the integration of previous locus‐specific databases for F7 into a single curated database with enhanced features. The database provides access to in silico analyses that may be useful in the prediction of variant pathogenicity as well as cross‐species sequence alignments, structural information, and functional and clinical severity described for each variant, where appropriate. The variant data is shared with the F7 Leiden Open Variation Database. The updated database now includes 221 unique variants, representing gene variants identified in 728 individuals. Single nucleotide variants are the most common type (88%) with missense representing 74% of these variants. A number of variants are found with relatively high minor allele frequencies that are not pathogenic but contribute significantly to the likely pathogenicity of coinherited variants due to their effect on FVII plasma levels. This comprehensive collection of curated information significantly aids the assessment of pathogenicity. [ABSTRACT FROM AUTHOR]

Introduction: Advances in genomic sequencing have facilitated the sequencing of genes associated with disorders of haemostasis. The identification of variants within genes and access to curated data incorporating structural, functional, evolutionary as well as phenotypic data has become increasingly important in order to ascribe pathogenicity. Aim: The European Association for Haemophilia and Allied Disorders (EAHAD) Coagulation Factor Variant Database Project aims to provide a single port of entry to a web‐accessible resource for variants in genes involved in clinical bleeding disorders. Results: New databases have evolved from previously developed single gene variant coagulation database projects, incorporating new data, new analysis tools and a new common database architecture with new interfaces and filters. These new databases currently present information about the genotype, phenotype (laboratory and clinical) and structural and functional effects of variants described in the genes of factor (F) VII (F7), FVIII (F8), FIX (F9) and von Willebrand factor (VWF). Conclusion: The project has improved the quality and quantity of information available to the haemostasis research and clinical communities, thereby enabling accurate classification of disease severity in order to make assessments of likely pathogenicity. [ABSTRACT FROM AUTHOR]