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The higher-order structure (HOS) of proteins plays a critical role in their function; therefore, it is important to our understanding of their function that we have as much information as possible about their three-dimensional structure and how it changes with time. Mass spectrometry (MS) has become an important tool for determining protein HOS owing to its high throughput, mid-to-high spatial resolution, low sample amount requirement and broad compatibility with various protein systems. Modern MS-based protein HOS analysis relies, in part, on footprinting, where a reagent reacts ‘to mark’ the solvent-accessible surface of the protein, and MS-enabled proteomic analysis locates the modifications to afford a footprint. Fast photochemical oxidation of proteins (FPOP), first introduced in 2005, has become a powerful approach for protein footprinting. Laser-induced hydrogen peroxide photolysis generates hydroxyl radicals that react with solvent-accessible side chains (14 out of 20 amino acid side chains) to fulfill the footprinting. The reaction takes place at sub-milliseconds, faster than most of labeling-induced protein conformational changes, thus enabling a ‘snapshot’ of protein HOS in solution. As a result, FPOP has been employed in solving several important problems, including mapping epitopes, following protein aggregation, locating small molecule binding, measuring ligand-binding affinity, monitoring protein folding and unfolding and determining hidden conformational changes invisible to other methods. Broader adoption will be promoted by dissemination of the technical details for assembling the FPOP platform and for dealing with the complexities of analyzing FPOP data. In this protocol, we describe the FPOP platform, the conditions for successful footprinting and its examination by mass measurements of the intact protein, the post-labeling sample handling and digestion, the liquid chromatography–tandem MS analysis of the digested sample and the data analysis with Protein Metrics Suite. This protocol is intended not only as a guide for investigators trying to establish an FPOP platform in their own lab but also for those willing to incorporate FPOP as an additional tool in addressing their questions of interest. Footprinting reports solvent accessibilities of a protein or protein complex. This protocol describes a fast photochemical oxidation of proteins platform for footprinting, with laser-initiated H2O2 photolysis and mass spectrometry analysis.

© 2018, Springer Science+Business Media, LLC, part of Springer Nature. The 24-item Five Facet Mindfulness Questionnaire – Short Form (FFMQ-SF) was developed to measure five facets of dispositional mindfulness: observing, describing, acting with awareness, non-judgment of inner experience, and non-reactivity to inner experience. The FFMQ-SF is increasingly used with older adult populations, despite not having been appropriately validated for such use. The present study examined the psychometric properties of the FFMQ-SF among community-dwelling older adults (N = 210). The five subscales of the FFMQ-SF were found to be internally consistent. Convergent validity analyses revealed that all facets of mindfulness except observing were negatively correlated with negative affect, and all facets except for non-judgment were positively correlated with positive affect. Similarly, all facets except for non-judgment were positively correlated with cognitive reappraisal emotion regulation tendencies. Only describing, acting with awareness, and non-judgment were negatively correlated with expressive suppression tendencies. The original five-factor structure of the FFMQ-SF was supported by confirmatory factor analyses. The two-factor higher order structure of the FFMQ-SF was also supported. Overall, the results support the psychometric properties of the FFMQ-SF for use with older adult samples.

Biologic products encounter various types of interfacial stress during development, manufacturing, and clinical administration. When proteins come in contact with vapor-liquid, solid-liquid, and liquid-liquid surfaces, these interfaces can significantly impact the protein drug product quality attributes, including formation of visible particles, subvisible particles, or soluble aggregates, or changes in target protein concentration due to adsorption of the molecule to various interfaces. Protein aggregation at interfaces is often accompanied by changes in conformation, as proteins modify their higher order structure in response to interfacial stresses such as hydrophobicity, charge, and mechanical stress. Formation of aggregates may elicit immunogenicity concerns; therefore, it is important to minimize opportunities for aggregation by performing a systematic evaluation of interfacial stress throughout the product development cycle and to develop appropriate mitigation strategies. The purpose of this white paper is to provide an understanding of protein interfacial stability, explore methods to understand interfacial behavior of proteins, then describe current industry approaches to address interfacial stability concerns. Specifically, we will discuss interfacial stresses to which proteins are exposed from drug substance manufacture through clinical administration, as well as the analytical techniques used to evaluate the resulting impact on the stability of the protein. A high-level mechanistic understanding of the relationship between interfacial stress and aggregation will be introduced, as well as some novel techniques for measuring and better understanding the interfacial behavior of proteins. Finally, some best practices in the evaluation and minimization of interfacial stress will be recommended.

Bacterial division begins with the formation of a contractile protein ring at midcell, which constricts the bacterial envelope to generate two daughter cells. The central component of the division ring is FtsZ, a tubulin-like protein capable of self-assembling into filaments which further associate into a higher order structure known as the Z ring. Proteins that bind to FtsZ play a crucial role in the formation and regulation of the Z ring. One such protein is ZapA, a widely conserved 21 kDa homodimeric protein that associates with FtsZ filaments and promotes their bundling. Although ZapA was discovered more than a decade ago, the structural details of its interaction with FtsZ remain unknown. In this work, backbone and side chain NMR assignments for the Geobacillus stearothermophilus ZapA homodimer are described. We titrated FtsZ into (15)N(2)H-ZapA and mapped ZapA residues whose resonances are perturbed upon FtsZ binding. This information provides a structural understanding of the interaction between FtsZ and ZapA.

The interphase nucleus and nuclear envelope can acquire a myriad of shapes in normal or pathological cell states. There exist a wide variety of indentations and invaginations, of protrusions and evaginations. It has been difficult to classify and name all of these nuclear shapes and, consequently, a barrier to understanding the biochemical and biophysical causes. This review focuses upon one type of nuclear envelope shape change, named "nuclear envelope-limited chromatin sheets" (ELCS), which appears to involve exaggerated nuclear envelope growth, carrying with it one or more layers of approximately 30 nm diameter heterochromatin. A hypothesis on the formation of ELCS is proposed, relating higher order heterochromatin structure in an interphase nucleus, nuclear envelope growth, and nuclear envelope-heterochromatin interactions.

A biosemiotic view of living things is presented that supersedes the mechanistic view of life prevalent in biology today. Living things are active agents with autonomous subjectivity, whose structure is triadic, consisting of the individual organism, its Umwelt and the society. Sociality inheres in every living thing since the very origin of life on the earth. The temporality of living things is guided by the purpose to live, which works as the semantic boundary condition for the processes of embodiment of the subjectivity. Freedom at the molecular and cellular levels allows autonomy and spontaneity to emerge even in single cell organisms, and the presence of the dimension of mind in every living thing is deduced. Living things transcend their individualness, as they live in historically formed higher order structure consisting of the lineage-species and the society. They also transcend materiality, having the dimension of mind.

The formation mechanism of banded spherulites of poly(vinylidene fluoride) has been examined experimentally by optical and atomic force microscopies. We have confirmed the proportional relationship between the band spacing and the maximum lateral width of lamellar crystals at the growth front, and the square root dependence of the maximum width and the band spacing on the growth rate. The square root dependence suggests a splaying instability of the growth front regulated by a gradient of free energy in the liquid side, and the proportional dependence suggests the dominant effect of torsional re-orientation on the occasion of splaying in the period determination of band spacing.

A major barrier to successful retrieval from external sources (e.g., electronic databases) is the tremendous variability in the words that people use to describe objects of interest. The fact that different authors use different words to describe essentially the same idea means that relevant objects will be missed; conversely, the fact that the same word can be used to refer to many different things means that irrelevant objects will be retrieved. We describe a statistical method called latent semantic indexing, which models the implicit higher order structure in the association of words and objects and improves retrieval performance by up to 30%. Additional large performance improvements of 40% and 67% can be achieved through the use of differential term weighting and iterative retrieval methods.

Large-scale mapping of chromatin state and transcription factor binding have uncovered many broad chromatin domains along linear genomic DNA, but it is unclear how these functional domains are organized in three-dimensional nuclear space. A new study now shows that many domains exist as loops connected by CCTC-binding factor (CTCF), providing new insights into the higher-order structure of chromatin organization in the nucleus.

In the north central zone of the Damara Orogen (Namibia) a field study was made of ptygmatic fold structures composed of more than one order of folds or cuspate interface structures of compressive origin. Their evolution can be described in terms of sequential development of fold orders. Each fold order is considered as a geometrically distinct group of fold structures, for it is impossible to predict the exact geometry of a higher order structure from the shape of a lower order fold or fold group.

The chromocenter heterochromatin of cell nuclei in the Scrophulariacean plant,Rhinanthus minor, was reconstructed from ultrathin serial sections by various methods. The chromocenters are irregularly shaped blocks through which runs a ramified anastomosing system of channels. No higher order structure of chromatin organization could be recognized.