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Evaluating the Effects of Hinge Flexibility on the Solution Structure of Antibodies at Concentrated Conditions.

ABSTRACT: Employing 2 different coarse-grained models, we evaluated the effect of intramolecular domain-domain distances and hinge flexibility on the general solution structure of monoclonal antibodies (mAbs), within the context of protein-protein steric repulsion. These models explicitly account for the hinge region, and represent antibodies at either domain or subdomain levels (i.e., 4-bead and 7-bead representations, respectively). Additionally, different levels of mAb flexibility are also considered. When evaluating mAbs as rigid structures, analysis of small-angle scattering profiles showed that changes in the relative internal distances between Fc and Fab domains significantly alter the local arrangement of neighboring molecules, as well as the molecular packing of the concentrated mAb solutions. Likewise, enabling hinge flexibility in either of the mAb models led to qualitatively similar results, where flexibility increases the spatial molecular arrangement at elevated concentrations. This occurs because fluctuations in mAb quaternary structure are modulated by the close proximity between molecules at elevated concentrations (>50 mg mL^-1), yielding an increased molecular packing and osmotic compressibility. However, our results also showed that the mechanism behind this synergy between flexibility and packing strongly depends on both the level of structural detail and the number of degrees-of-freedom considered in the coarse-grained model.

SUBMITTER: Blanco MA

PROVIDER: S-EPMC7288351 | biostudies-literature | 2019 May

REPOSITORIES: biostudies-literature

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Evaluating the Effects of Hinge Flexibility on the Solution Structure of Antibodies at Concentrated Conditions.

Blanco Marco A MA Hatch Harold W HW Curtis Joseph E JE Shen Vincent K VK

Journal of pharmaceutical sciences 20181226 5

Employing 2 different coarse-grained models, we evaluated the effect of intramolecular domain-domain distances and hinge flexibility on the general solution structure of monoclonal antibodies (mAbs), within the context of protein-protein steric repulsion. These models explicitly account for the hinge region, and represent antibodies at either domain or subdomain levels (i.e., 4-bead and 7-bead representations, respectively). Additionally, different levels of mAb flexibility are also considered. ...[more]

PMID: 30593783

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Project description:The interplay between environmental and genetic factors conditions the fruit ripening program in plants. Transcriptome analysis of grapevine fruits can help understanding these interactions to consciously cope with conditions leading to detrimental effects for viticultural purposes. However, considering the grapevine characteristic ripening asynchrony, which can be intensified by contrasting conditions, accurate grape sampling may be essential for molecular comparisons. In this study, berry density sorting according to floatability in NaCl solutions was assessed as a grape ripening staging strategy. Total sugar content was more correlated with berry density than with other non-invasive ripening parameters. The transcriptome was compared between three density classes collected near commercial maturity using grapevine whole-genome NimbleGen microarrays. Expression profiles clearly related with ripening progression were detected in a density series simultaneously collected from a vineyard of Albariño. By contrast, considerable differences were detected when the same density series was sampled on two different dates from the same vineyard of Tempranillo. Functional analysis indicated that environmental differences between both sampling moments determined most of these expression differences. Ripening degree-dependent responses to the environment were also detected. Finally, the effect of the sorting procedures on the grape transcriptome showed negligible when it was directly tested. Altogether, these findings evidence the convenience of homogenizing the developmental stage and the sampling time conditions for transcriptome comparisons. Berry density sorting proved useful to this end, although this method could be limited when the berry sugar concentration is not determined by the ripening developmental program. The potential berry transcriptional response to the high concentrated salt solutions used for berry density-sorting may involve a concern on the suitability of this method for transcriptomic studies. This possibility was directly tested by comparing the expression of berries immersed in a highly concentrated NaCl solution (170 g•L-1) during a relatively long time lapse (1 h) with that in berries kept at room air. This experiment was carried out using Tempranillo grapes at mid-véraison to standardize the ripening state by selecting berries with the same proportion of coloured skin surface (50%, corresponding to TSS of 9.9 ±0.3 ºBrix). Berries grown under watering (W) and no-watering (NW) conditions were analysed to test for possible irrigation-NaCl solution interactions affecting berry gene expression. No significant DEG was identified when the effect of the NaCl solution was analysed separately for berries collected from W or NW blocks (≥2-fold change and B-H adjusted P≤0.05 in 2-class limma). When W and NW samples were used together to analyse the effect of the NaCl immersion irrespectively of the irrigation treatment, 15 genes were detected with a B-H adjusted P≤0.05 and only 13 of these showed ≥2-fold change, confirming the slight effect. In summary, the effect of the NaCl immersion on the berry transcriptome is negligible and does not seem to interact with the transcriptomic response to other environmental cues.

Dataset Information

Evaluating the Effects of Hinge Flexibility on the Solution Structure of Antibodies at Concentrated Conditions.

Publications

Evaluating the Effects of Hinge Flexibility on the Solution Structure of Antibodies at Concentrated Conditions.

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