Efforts to improve detection sensitivity for capillary electrophoresis coupled to atmospheric pressure photoionization mass spectrometry

Electrospray ionization performs best with volatile buffers. However, generally the best separation performance for capillary electrophoresis (CE) is achieved with non‐volatile buffers. Hyphenation of CE with mass spectrometry (MS) utilizing atmospheric pressure photoionization (APPI) enables use of a wider range of separation buffers without compromising detection sensitivity. As APPI is considered to be mass flow sensitive, the use of a larger inner diameter separation capillary (75 µm) allows larger volumes to be injected, without decreased separation performance, thus providing improved sensitivity (approx. a factor of 10), compared to the use of a 25 µm capillary. However, nebulizing gas flow and position of capillary tip in the sprayer have to be carefully optimized to prevent excessive band broadening. Further improvement in sensitivity (approx. a factor of 2) was obtained by decreasing the distance between the sprayer and ionization region, indicating that a specially designed CE/APPI‐MS interface for low flow rates will be favourable. Copyright © 2010 John Wiley & Sons, Ltd.     

Local cooperativity in an amyloidogenic state of human lysozyme observed at atomic resolution

The partial unfolding of human lysozyme underlies its conversion from the soluble state into amyloid fibrils observed in a fatal hereditary form of systemic amyloidosis. To understand the molecular origins of the disease, it is critical to characterize the structural and physicochemical properties of the amyloidogenic states of the protein. Here we provide a high-resolution view of the unfolding process at low pH for three different lysozyme variants, the wild-type protein and the mutants I56T and I59T, which show variable stabilities and propensities to aggregate in vitro. Using a range of biophysical techniques that includes differential scanning calorimetry and nuclear magnetic resonance spectroscopy, we demonstrate that thermal unfolding under amyloidogenic solution conditions involves a cooperative loss of native tertiary structure, followed by progressive unfolding of a compact, molten globule-like denatured state ensemble as the temperature is increased. The width of the temperature window over which the denatured ensemble progressively unfolds correlates with the relative amyloidogenicity and stability of these variants, and the region of lysozyme that unfolds first maps to that which forms the core of the amyloid fibrils formed under similar conditions. Together, these results present a coherent picture at atomic resolution of the initial events underlying amyloid formation by a globular protein.     

Optimizing the performance of tin dioxide microspheres for phosphopeptide enrichment

Phosphopeptide enrichment based on metal oxide affinity chromatography is one of the most powerful tools for studying protein phosphorylation on a large scale. To complement existing metal oxide sorbents, we have recently introduced tin dioxide as a promising alternative. The preparation of SnO₂ microspheres by the nanocasting technique, using silica of different morphology as a template, offers a strategy to prepare materials that vary in their particle size and their porosity. Here, we demonstrate how such stannia materials can be successfully generated and their properties fine-tuned in order to obtain an optimized phosphopeptide enrichment material. We combined data from liquid chromatography-mass spectrometry experiments and physicochemical characterization, including nitrogen physisorption and energy-dispersive X-ray spectroscopy (EDX), to explain the influence of the various experimental parameters.     

Dirac structures and their composition on Hilbert spaces

Dirac structures appear naturally in the study of certain classes of physical models described by partial differential equations and they can be regarded as the underlying power conserving structures. We study these structures and their properties from an operator-theoretic point of view. In particular, we find necessary and sufficient conditions for the composition of two Dirac structures to be a Dirac structure and we show that they can be seen as Lagrangian (hyper-maximal neutral) subspaces of Kre?n spaces. Moreover, special emphasis is laid on Dirac structures associated with operator colligations. It turns out that this class of Dirac structures is linked to boundary triplets and that this class is closed under composition.     

Well-Posed State/Signal Systems in Continuous Time

We introduce a new class of linear systems, the L ^p -well-posed state/signal systems in continuous time, we establish the foundations of their theory and we develop some tools for their study. The principal feature of a state/signal system is that the external signals of the system are not a priori divided into inputs and outputs. We relate state/signal systems to the better-known class of well-posed input/state/output systems, showing that state/signal systems are more flexible than input/state/output systems but still have enough structure to provide a meaningful theory. We also give some examples which point to possibilities for further study.