Fluorogenic probes for detecting deacylase and demethylase activity towards post-translationally-modified lysine residues

Reversible enzymatic post-translational modification of the ε-amino groups of lysine residues (e.g. N-acylation reactions) plays an important role in regulating the cellular activities of numerous proteins. This study describes how enzyme catalyzed N-deprotection of lysine residues of non-fluorescent peptide-coumarin probes can be used to generate N-deprotected peptides that undergo spontaneous O- to N-ester transfer reactions (uncatalyzed) to generate a highly fluorescent N-carbamoyl peptide. This enables detection of enzyme catalyzed N-deacetylation, N-demalonylation, N-desuccinylation and N-demethylation reactions activities towards the N-modified lysine residues of these probes using simple ‘turn on’ fluorescent assays.

We developed “turn-on” fluorescent probes that detect enzymatic lysine deacylation and demethylation critical for epigenetic and other cellular phenomena, using intramolecular O- to N-ester transfer reactions.     

Tuning cell adhesion on gradient poly(2-hydroxyethyl methacrylate)-grafted surfaces

A simple yet versatile method was developed to prepare a low-density polymerization initiator gradient, which was combined with surface-initiated atom transfer radical polymerization (ATRP) to produce a well-defined poly(2-hydroxyethyl methacrylate) (HEMA) gradient substrate. A smooth variation in film thickness was measured across the gradient, ranging from 20 A to over 80 A, but we observed a nonmonotonic variation in water contact angle. Fits of X-ray reflectivity profiles suggested that at the low graft density end, the polymer chain structure was in a "mushroom" regime, while the polymer chains at high graft density were in a "brush" regime. It was found that the "mushroom" region of the gradient could be made adhesive to cells by adsorbing adhesion proteins, and cell adhesion could be tuned by controlling the density of the polymer grafts. Fibroblasts were seeded on gradients precoated with fibronectin to test cellular responses to this novel substrate, but it was found that cell adhesion did not follow the expected trend; instead, saturated cell adhesion and spreading was found at the low grafting density region.     

Liquid chromatographic studies of the effect of phosphate on the binding properties of silica-immobilized bovine serum albumin

High-performance liquid chromatography has been used to examine how phosphate ions affect the binding properties of bovine serum albumin (BSA) immobilized to porous silica. In doing this, the time dependence of the protein to reach conformational equilibrium is measured as a function of the concentration of phosphate in the eluent using the D- and L-isomers of tryptophan and kynurenine as solutes. The overall binding and chiral selectivity (alphaD,L) of the protein toward these solutes appear to be related to two types of effects: one being those that are site-selective and only influence the retention of the L-isomers and the other being those that are nonselective and influence the retention of both enantiomers. An interesting feature of the concentration-dependent data is a maximum in alphaD,L at intermediate phosphate concentrations (i.e., 10 to 50mM phosphate) indicative of both cooperative and antagonistic binding effects. Phosphate eluents within this concentration range provide selectivity advantages, and those at higher concentrations decrease the time required for the protein or column to reach equilibrium. A final set of studies has also been carried out using four alternate buffer systems (i.e., borate, carbonate, acetate, and arsenate eluents). Although the borate eluents affect the BSA's binding properties and alphaD,L similar to the phosphate eluents, the other buffers result in poor separations. Observations from this study are useful in helping to optimize separations carried out on immobilized BSA as well as addressing biological and mechanistic questions related to how anions influence the native binding properties of serum albumins.     

Effect of Ce<Superscript>3+</Superscript> on spectral characteristic of D1/D2/Cytb559 complex from spinach

It was studied by spectroscopy that PSII reaction center complex consisting of three polypeptides, D₁, D₂ and Cytb559, were purified from PSII particle of CeCl₃ treated spinach. The results of the experiment show that Ce³⁺ could improve the growth of spinach, and accelerate electron transport of PSII particles. Of chl-a of UV-Vis spectrum of D1/D2/Cytb559 complex, Soret band was blue-shifted by 3 nm and Q band by 2 nm, respectively, and the fluorescence emission peak was blue-shifted by 5 nm in CeCl₃-treated spinach compared with the one in control. By the extended X-ray absorption fine structure (EXAFS) spectroscopy methods, it has been found that Ce³⁺ is coordinated with 8 nitrogen atoms in the first coordination shell with Ce-N bond length of 0.253 nm, and Ce³⁺ with 6 oxygen atoms in the second coordination shell with Ce-O bond length of 0.32 nm. However, the secondary structure of D1/D2/Cytb559 complex by circular dichroism (CD) spectroscopy has no significant change after CeCl₃ treated. It might be that Ce³⁺ binds to porphyrin rings of chlorophyll and oxygen of amino acid residue of polypeptide in D1/D2/Cytb559 complex, and then accelerates the primary reaction of PSII, intensifies function of P680⁺ primary electron donor of D1/D2/Cytb559, but there is little change in conformation of PSII reaction center complex.     

On the integrability of a four-component super system

Theoretical and Mathematical Physics - A four-component system of the Camassa–Holm type depending on both bosonic and fermionic variables is proposed. This system is shown to be the...     

Experimental study of natural convection heat transfer between an outer horizontal cylindrical envelope and an inner concentric heated square cylinder with two slots

This paper presents the results of an experimental study of natural convection heat transfer between a horizontal cylindrical envelope and an internal concentric heated square cylinder with two slots. The internal cylinder was a hollow one with horizontal slots on its top and bottom surfaces. The ratio of slot widthS to the side heightH was 0.0612 and 0.3878. The ratio of the envelope inner diameterD _o to the side heightH was 2.653. Air was used as the working fluid. The range of Ray-leigh number was 1.77×10²8.72×10⁶ forS/H=0.0612 and 1.32×10²6.25×10⁶ forS/H=0.3878. The results show that there are three different heat transfer regimes in different Ray-leigh number regions, i.e. pure conduction regime, transition regime and convection regime. The average heat transfer results were correlated into two empirical equations. Comparison was made with the non-slotted case. It is found that slots of the internal cylinder can significantly enhance the heat transfer.

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Experimentelle Untersuchung des Wärmeübergangs bei natürlicher Konvektion zwischen einer horizontalen zylindrischen Außenhülle und einem konzentrischen, beheizten, quadratischen Prisma mit zwei Schlitzen

Zusammenfassung In der Arbeit werden die Ergebnisse einer experimentellen Untersuchung des Wärmeübergangs bei natürlicher Konvektion zwischen einer horizontalen zylindrischen Außenhülle und einem beheizten quadratischen Prisma mit zwei Schlitzen vorgestellt. Das Prisma selbst ist hohl und weist in der oberen und unteren Begrenzungsfläche je einen horizontalen Längsschlitz auf. Das Verhältnis von SchlitzweiteS zu SeitenhöheH beträgt 0,0612 und 0,3878, das des HülleninnendurchmessersD _o zur SeitenhöheH beträgt 2,653. Als Arbeitsmedium diente Luft. Die Rayleigh-Zahlen variierten zwischen 1,7·10² und 8,72·10⁶ fürS/H=0,0612 und zwischen 1,32·10² und 6,25·10⁶ fürS/H=0,3878. Die Ergebnisse belegen die Existenz dreier unterschiedlicher Wärmeübergangsregime in den verschiedenen Rayleigh-Zahl-Bereichen, und zwar reiner Leitungsbereich, Übergangsgebiet und Konvektionsbereich. Die Ergebnisse für den Wärmeübergang werden im Vergleich mit jenen für ein Prisma ohne Schlitze durch zwei Korrelationbeziehungen dargestellt. Es zeigt sich, daß durch Anbringung von Schlitzen am Innenprisma der Wärmeübergang wesentlich verstärkt werden kann.

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Nomenclature C _p specific heat at constant pressure, J/(kg·K) - D _i diameter of the related circular cylinder whose circumferential area is equal to that of the unslotted square cylinder, m - D _o internal diameter of the outer circular envelope, m - F _i surface area of the inner two slot cylinder, m - g gravitational acceleration, m/s² - H distance between the opposite sides of the square cylinder with two slots, m - K _eq dimensionless equivalent thermal conductivity - L axial length of the test section, m - m ratio of the area of the unslotted square cylinder surface to that of the slotted square cylinder - P pressure in the enclosure, Pa - Q total power input to the enclosure, W - Q _cond radial heat conduction, W - Q _conv convective heat transfer, W - Q _r radiation heat transfer, W - Q _los end heat dissipation, W - R air gas constant, J/(kg·K) - Ra Rayleigh number - S slot width, m - T _i wall temperature of the inner cylinder, K - T _o wall temperature of the outer envelope, K - T _m mean temperature, K - T temperature difference=T _i –T _o , K - W maximum gap width of the test annuli=(D _o –H)/2 for the square case, m Greek symbols ₀ black body radiation constant, W/(m²·K⁴) - _s equation system emissivity - air thermal conductivity, W/(m·K) - _eq equivalent thermal conductivity, W/(m·K) - air dynamic viscosity, kg/(m·s) This work was supported by the National Natural Science Foundation of China.