Atomistic Modeling of Scattering Curves for Human IgG1/4 Reveals New Structure-Function Insights

Small angle x-ray and neutron scattering are techniques that give solution structures for large macromolecules. The creation of physically realistic atomistic models from known high-resolution structures to determine joint x-ray and neutron scattering best-fit structures offers a, to our knowledge, new method that significantly enhances the utility of scattering. To validate this approach, we determined scattering curves for two human antibody subclasses, immunoglobulin G (IgG) 1 and IgG4, on five different x-ray and neutron instruments to show that these were reproducible, then we modeled these by Monte Carlo simulations. The two antibodies have different hinge lengths that connect their antigen-binding Fab and effector-binding Fc regions. Starting from 231,492 and 190,437 acceptable conformations for IgG1 and IgG4, respectively, joint x-ray and neutron scattering curve fits gave low goodness-of-fit R factors for 28 IgG1 and 2748 IgG4 structures that satisfied the disulphide connectivity in their hinges. These joint best-fit structures showed that the best-fit IgG1 models had a greater separation between the centers of their Fab regions than those for IgG4, in agreement with their hinge lengths of 15 and 12 residues, respectively. The resulting asymmetric IgG1 solution structures resembled its crystal structure. Both symmetric and asymmetric solution structures were determined for IgG4. Docking simulations with our best-fit IgG4 structures showed greater steric clashes with its receptor to explain its weaker FcγRI receptor binding compared to our best-fit IgG1 structures with fewer clashes and stronger receptor binding. Compared to earlier approaches for fitting molecular antibody structures by solution scattering, we conclude that this joint fit approach based on x-ray and neutron scattering data, combined with Monte Carlo simulations, significantly improved our understanding of antibody solution structures. The atomistic nature of the output extended our understanding of known functional differences in Fc receptor binding between IgG1 and IgG4

The Fab conformations in the solution structure of human IgG4 restricts access to its Fc region: implications for functional activity.

Human IgG4 antibody shows therapeutically-useful properties compared to the IgG1, IgG2 and IgG3 subclasses. Thus IgG4 does not activate complement, and shows conformational variability. These properties are attributable to its hinge region, which is the shortest of the four IgG subclasses. Using high throughput scattering methods, we have studied the solution structure of wild-type IgG4(Ser222) and a hinge mutant IgG4(Pro222) in different buffers and temperatures, where the proline substitution suppresses the formation of half-antibody. Analytical ultracentrifugation showed that both IgG4 forms were principally monomeric with sedimentation coefficients s020,w of 6.6-6.8 S. A monomer-dimer equilibrium was observed in heavy water buffer at low temperature. Scattering showed that the X-ray radius of gyration RG was unchanged with concentration in 50-250 mM NaCl buffers, while the neutron RG values showed a concentration-dependent increase as the temperature decreased in heavy water buffers. The distance distribution curves P(r) revealed two peaks, M1 and M2 that shifted below 2 mg/ml to indicate concentration-dependent IgG4 structures, in addition to IgG4 dimer formation at high concentration in heavy water. Constrained X-ray and neutron scattering modelling revealed asymmetric solution structures for IgG4(Ser222) with extended hinge structures. The IgG4(Pro222) structure was similar. Both IgG4 structures showed that their Fab regions were positioned close enough to the Fc region to restrict C1q binding. Our new molecular models for IgG4 explain its inability to activate complement, and clarifies aspects of its stability and function for therapeutic applications

The solution structure of the human IgG2 subclass is distinct from those for human IgG1 and IgG4 providing an explanation for their discrete functions

Human IgG2 antibody displays distinct therapeutically-useful properties compared with the IgG1, IgG3 and IgG4 antibody subclasses. IgG2 is the second most abundant IgG subclass, being able to bind human FcγRII/FcγRIII, but not to FcγRI or complement C1q. Structural information on IgG2 is limited by the absence of a full-length crystal structure for this. To this end, we determined the solution structure of human myeloma IgG2 by atomistic X-ray and neutron scattering modelling. Analytical ultracentrifugation disclosed that IgG2 is monomeric with a sedimentation coefficient s020,w of 7.2 S. IgG2 dimer formation was ≤ 5% and independent of the buffer conditions. Small-angle X-ray scattering in a range of NaCl concentrations and in light and heavy water revealed that the X-ray radius of gyration Rg is 5.2-5.4 nm, after allowing for radiation damage at higher concentrations, and that the neutron Rg value of 5.0 nm remained unchanged in all conditions. The X-ray and neutron distance distribution curves P(r) revealed two peaks, M1 and M2, that were unchanged in different buffers. The creation of ˃123,000 physically-realistic atomistic models by Monte Carlo simulations for joint X-ray and neutron-scattering curve fits, constrained by the requirement of correct disulfide bridges in the hinge, resulted in the determination of symmetric Y-shaped IgG2 structures. These molecular structures were distinct from those for asymmetric IgG1 and asymmetric and symmetric IgG4, and were attributable to the four hinge disulfides. Our IgG2 structures rationalize the existence of the human IgG1, IgG2, and IgG4 subclasses, and explain the receptor binding functions of IgG2

Ultrathin compound semiconductor on insulator layers for high performance nanoscale transistors

Over the past several years, the inherent scaling limitations of electron devices have fueled the exploration of high carrier mobility semiconductors as a Si replacement to further enhance the device performance. In particular, compound semiconductors heterogeneously integrated on Si substrates have been actively studied, combining the high mobility of III-V semiconductors and the well-established, low cost processing of Si technology. This integration, however, presents significant challenges. Conventionally, heteroepitaxial growth of complex multilayers on Si has been explored. Besides complexity, high defect densities and junction leakage currents present limitations in the approach. Motivated by this challenge, here we utilize an epitaxial transfer method for the integration of ultrathin layers of single-crystalline InAs on Si/SiO2 substrates. As a parallel to silicon-on-insulator (SOI) technology14,we use the abbreviation "XOI" to represent our compound semiconductor-on-insulator platform. Through experiments and simulation, the electrical properties of InAs XOI transistors are explored, elucidating the critical role of quantum confinement in the transport properties of ultrathin XOI layers. Importantly, a high quality InAs/dielectric interface is obtained by the use of a novel thermally grown interfacial InAsOx layer (~1 nm thick). The fabricated FETs exhibit an impressive peak transconductance of ~1.6 mS/{\mu}m at VDS=0.5V with ON/OFF current ratio of greater than 10,000 and a subthreshold swing of 107-150 mV/decade for a channel length of ~0.5 {\mu}m

Gene products of chromosome 11q and their association with CCND1 gene amplification and tamoxifen resistance in premenopausal breast cancer

Introduction: The amplification event occurring at chromosome locus 11q13, reported in several different cancers, includes a number of potential oncogenes. We have previously reported amplification of one such oncogene, namely CCND1, to be correlated with an adverse effect of tamoxifen in premenopausal breast cancer patients. Over-expression of cyclin D-1 protein, however, confers tamoxifen resistance but not a tamoxifen-induced adverse effect. Potentially, co-amplification of an additional 11q13 gene, with a resulting protein over-expression, is required to cause an agonistic effect. Moreover, during 11q13 amplification a deletion of the distal 11q region has been described. In order to assess the potential impact of the deletion we examined a selected marker for this event. Method: Array comparative genomic hybridization analysis was employed to identify and confirm changes in the gene expression of a number of different genes mapping to the 11q chromosomal region, associated with CCND1 amplification. The subsequent protein expression of these candidate genes was then examined in a clinical material of 500 primary breast cancers from premenopausal patients who were randomly assigned to either tamoxifen or no adjuvant treatment. The protein expression was also compared with gene expression data in a subset of 56 breast cancer samples. Results: Cortactin and FADD (Fas-associated death domain) over-expression was linked to CCND1 amplification, determined by fluorescence in situ hybridization, but was not associated with a diminished effect of tamoxifen. However, deletion of distal chromosome 11q, defined as downregulation of the marker Chk1 (checkpoint kinase 1), was associated with an impaired tamoxifen response, and interestingly with low proliferative breast cancer of low grade. For Pak1 (p21-activated kinase 1) and cyclin D-1 the protein expression corresponded to the gene expression data. Conclusions: The results indicate that many 11q13 associated gene products are over-expressed in conjunction with cyclin D-1 but not linked to an agonistic effect of tamoxifen. Finally, the deletion of distal 11q, linked to 11q13 amplification, might be an important event affecting breast cancer outcome and tamoxifen response

Hollow Sodium Tungsten Bronze (Na0.15WO3) Nanospheres: Preparation, Characterization, and Their Adsorption Properties

We report herein a facile method for the preparation of sodium tungsten bronzes hollow nanospheres using hydrogen gas bubbles as reactant for chemical reduction of tungstate to tungsten and as template for the formation of hollow nanospheres at the same time. The chemical composition and the crystalline state of the as-prepared hollow Na0.15WO3nanospheres were characterized complementarily, and the hollow structure formation mechanism was proposed. The hollow Na0.15WO3nanospheres showed large Brunauer–Emment–Teller specific area (33.8 m2 g−1), strong resistance to acids, and excellent ability to remove organic molecules such as dye and proteins from aqueous solutions. These illustrate that the hollow nanospheres of Na0.15WO3should be a useful adsorbent

Toward a multiscale modeling framework for understanding serotonergic function

Despite its importance in regulating emotion and mental wellbeing, the complex structure and function of the serotonergic system present formidable challenges toward understanding its mechanisms. In this paper, we review studies investigating the interactions between serotonergic and related brain systems and their behavior at multiple scales, with a focus on biologically-based computational modeling. We first discuss serotonergic intracellular signaling and neuronal excitability, followed by neuronal circuit and systems levels. At each level of organization, we will discuss the experimental work accompanied by related computational modeling work. We then suggest that a multiscale modeling approach that integrates the various levels of neurobiological organization could potentially transform the way we understand the complex functions associated with serotonin

Moxibustion for cancer care: a systematic review and meta-analysis

<p>Abstract</p> <p>Background</p> <p>Moxibustion is a traditional Chinese method that uses the heat generated by burning herbal preparations containing <it>Artemisia vulgaris </it>to stimulate acupuncture points. Considering moxibustion is closely related to acupuncture, it seems pertinent to evaluate the effectiveness of moxibustion as a treatment of symptoms of cancer. The objective of this review was to systematically assess the effectiveness of moxibustion for supportive cancer care.</p> <p>Methods</p> <p>We searched the literature using 11 databases from their inceptions to February 2010, without language restrictions. We included randomised clinical trials (RCTs) in which moxibustion was employed as an adjuvant treatment for conventional medicine in patients with any type of cancer. The selection of studies, data extraction, and validations were performed independently by two reviewers.</p> <p>Results</p> <p>Five RCTs compared the effects of moxibustion with conventional therapy. Four RCTs failed to show favourable effects of moxibustion for response rate compared with chemotherapy (n = 229, RR, 1.04, 95% CI 0.94 to 1.15, P = 0.43). Two RCTs assessed the occurrence of side effects of chemotherapy and showed favourable effects of moxibustion. A meta-analysis showed significant less frequency of nausea and vomiting from chemotherapy for moxibustion group (n = 80, RR, 0.38, 95% CIs 0.22 to 0.65, P = 0.0005, heterogeneity: χ²= 0.18, P = 0.67, I²= 0%).</p> <p>Conclusion</p> <p>The evidence is limited to suggest moxibustion is an effective supportive cancer care in nausea and vomiting. However, all studies have a high risk of bias so effectively there is not enough evidence to draw any conclusion. Further research is required to investigate whether there are specific benefits of moxibustion for supportive cancer care.</p

BRIT1/MCPH1 links chromatin remodelling to DNA damage response

To detect and repair damaged DNA, DNA damage response proteins need to overcome the barrier of condensed chromatin to gain access to DNA lesions1. ATP-dependent chromatin remodeling is one of the fundamental mechanisms used by cells to relax chromatin in DNA repair2–3. However, the mechanism mediating their recruitment to DNA lesions remains largely unknown. BRIT1 (also known as MCPH1) is an early DNA damage response protein that is mutated in human primary microcephaly4–8. We report here a previously unknown function of BRIT1 as a regulator of ATP-dependent chromatin remodeling complex SWI/SNF in DNA repair. Upon DNA damage, BRIT1 increases its interaction with SWI/SNF through the ATM/ATR-dependent phosphorylation on the BAF170 subunit. This increase of binding affinity provides a means by which SWI/SNF can be specifically recruited to and maintained at DNA lesions. Loss of BRIT1 causes impaired chromatin relaxation owing to reduced association of SWI/SNF with chromatin. This explains the decreased recruitment of repair proteins to DNA lesions and reduced efficiency of repair in BRIT1-deficient cells, resulting in impaired survival from DNA damage. Our findings, therefore, identify BRIT1 as a key molecule that links chromatin remodeling with DNA damage response in the control of DNA repair, and its dysfunction contributes to human disease