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Knowledge‐based methods for analyzing protein structures, such as statistical potentials, primarily consider the distances between pairs of bodies (atoms or groups of atoms). Considerations of several bodies simultaneously are generally used to characterize bonded structural elements or those in close contact with each other, but historically do not consider atoms that are not in direct contact with each other. In this report, we introduce an information‐theoretic method for detecting and quantifying distance‐dependent through‐space multibody relationships between the sidechains of three residues. The technique introduced is capable of producing convergent and consistent results when applied to a sufficiently large database of randomly chosen, experimentally solved protein structures. The results of our study can be shown to reproduce established physico‐chemical properties of residues as well as more recently discovered properties and interactions. These results offer insight into the numerous roles that residues play in protein structure, as well as relationships between residue function, protein structure, and evolution. The techniques and insights presented in this work should be useful in the future development of novel knowledge‐based tools for the evaluation of protein structure. Proteins 2014; 82:3450–3465. © 2014 Wiley Periodicals, Inc. 相似文献
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Mi Jin Nicolas Szapiel Jennifer Zhang John Hickey Sanchayita Ghose 《Biotechnology and bioengineering》2010,105(2):306-316
Host cell proteins (HCPs) constitute a major group of impurities for biologic drugs produced using cell culture technology. HCPs are required to be closely monitored and adequately removed in the downstream process. However, HCPs are a complex mixture of proteins with significantly diverse molecular and immunological properties. An overall understanding of the composition of HCPs and changes in their molecular properties upon changes in upstream and harvest process conditions can greatly facilitate downstream process design. This article describes the use of a comparative proteomic profiling method viz. two‐dimensional difference gel electrophoresis (2D‐DIGE) to examine HCP composition in the harvest stream of CHO cell culture. The effect of upstream process parameters such as cell culture media, bioreactor control strategy, feeding strategy, and cell culture duration/cell viability on HCP profile was examined using this technique. Among all the parameters studied, cell viability generated the most significant changes on the HCP profile. 2D‐DIGE was also used to compare the HCP differences between monoclonal antibody producing and null cell cultures. The HCP species in production cell culture was found to be well represented in null cell culture, which confirms the suitability of using the null cell culture for immunoassay reagent generation. 2D‐DIGE is complimentary to the commonly used HCP immunoassay. It provides a direct comparison of the changes in HCP composition under different conditions and can reveal properties (pI, MW) of individual species, whereas the immunoassay sensitively quantifies total HCP amount in a given sample. Biotechnol. Bioeng. 2010; 105: 306–316. © 2009 Wiley Periodicals, Inc. 相似文献
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Artificial proteins as allosteric modulators of PDZ3 and SH3 in two‐domain constructs: A computational characterization of novel chimeric proteins
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Palani Kirubakaran Lucie Pfeiferová Kristýna Boušová Lucie Bednarova Veronika Obšilová Jiří Vondrášek 《Proteins》2016,84(10):1358-1374
Artificial multidomain proteins with enhanced structural and functional properties can be utilized in a broad spectrum of applications. The design of chimeric fusion proteins utilizing protein domains or one‐domain miniproteins as building blocks is an important advancement for the creation of new biomolecules for biotechnology and medical applications. However, computational studies to describe in detail the dynamics and geometry properties of two‐domain constructs made from structurally and functionally different proteins are lacking. Here, we tested an in silico design strategy using all‐atom explicit solvent molecular dynamics simulations. The well‐characterized PDZ3 and SH3 domains of human zonula occludens (ZO‐1) (3TSZ), along with 5 artificial domains and 2 types of molecular linkers, were selected to construct chimeric two‐domain molecules. The influence of the artificial domains on the structure and dynamics of the PDZ3 and SH3 domains was determined using a range of analyses. We conclude that the artificial domains can function as allosteric modulators of the PDZ3 and SH3 domains. Proteins 2016; 84:1358–1374. © 2016 Wiley Periodicals, Inc. 相似文献
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Rapid one‐step purification of single‐cells encapsulated in alginate microcapsules from oil to aqueous phase using a hydrophobic filter paper: Implications for single‐cell experiments
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By virtue of the biocompatibility and physical properties of hydrogel, picoliter‐sized hydrogel microcapsules have been considered to be a biometric signature containing several features similar to that of encapsulated single cells, including phenotype, viability, and intracellular content. To maximize the experimental potential of encapsulating cells in hydrogel microcapsules, a method that enables efficient hydrogel microcapsule purification from oil is necessary. Current methods based on centrifugation for the conventional stepwise rinsing of oil, are slow and laborious and decrease the monodispersity and yield of the recovered hydrogel microcapsules. To remedy these shortcomings we have developed a simple one‐step method to purify alginate microcapsules, containing a single live cell, from oil to aqueous phase. This method employs oil impregnation using a commercially available hydrophobic filter paper without multistep centrifugal purification and complicated microchannel networks. The oil‐suspended alginate microcapsules encapsulating single cells from mammalian cancer cell lines (MCF–7, HepG2, and U937) and microorganisms (Chlorella vulgaris) were successfully exchanged to cell culture media by quick (~10 min) depletion of the surrounding oil phase without coalescence of neighboring microcapsules. Cell proliferation and high integrity of the microcapsules were also demonstrated by long‐term incubation of microcapsules containing a single live cell. We expect that this method for the simple and rapid purification of encapsulated single‐cell microcapsules will attain widespread adoption, assisting cell biologists and clinicians in the development of single‐cell experiments. 相似文献