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Spontaneous Cleavage (spontaneous + cleavage)
Selected AbstractsProduction of native and modified recombinant Der p 1 molecules in tobacco plantsCLINICAL & EXPERIMENTAL ALLERGY, Issue 5 2009D. Burtin Summary Background As a complex molecule requiring post-translational processing, it has been difficult to produce the Der p 1 major allergen from the Dermatophagoides pteronyssinus house dust mite in a recombinant form. Objective Here, we tested whether transgenic tobacco plants are suitable to express Der p 1, either as a wild-type molecule or as variants lacking N -glycosylation sites (Gly,) and/or cysteine protease activity (Enz,). Methods Using Agrobacterium tumefaciens -based transformation, pro Der p 1 molecules bearing mutations within either the N -glycosylation sites (N34Q, N150Q) and/or the cysteine protease-active site (C132V) were expressed in tobacco plants. After purification by ion exchange chromatography, allergens were characterized using immunoblotting, circular dichroism (CD), as well as basophil and T lymphocyte stimulation assays. Results Four forms of recombinant Der p 1 (i.e. wild-type Gly+/Enz+, as well as Gly,/Enz+, Gly+/Enz, or Gly,/Enz, variants) were successfully expressed in tobacco leaves as pro Der p 1 molecules. Spontaneous cleavage of the pro-peptide was observed in tobacco leaf extracts for all forms of recombinant Der p 1 (r Der p 1). CD confirmed that all r Der p 1 molecules, with the exception of the Gly,/Enz, variant, exhibited secondary structures comparable to the natural protein. A cysteine protease activity was associated only with the Gly+/Enz+ form. All these molecules exhibit a profile similar to natural Der p 1 with respect to IgE immunoreactivity, basophil activation and T cell recognition. Conclusion A tobacco plant expression system allows the production of various forms of mature Der p 1, which could be used for diagnostic or immunotherapeutic purposes. [source] Cleavage of CO by Mo[N(R)Ar]3 ComplexesEUROPEAN JOURNAL OF INORGANIC CHEMISTRY, Issue 23 2007Gemma Christian Abstract The reaction of MoL3 [L = NH2 and N(tBu)Ar] with CO was explored using DFT in order to rationalize why CO cleavage is not observed experimentally for this system in contrast to the corresponding N2 reaction which results in spontaneous cleavage of the N,N bond. The binding of CO to MoL3 was found to be both kinetically and thermodynamically favored over the binding of N2, with the formation of the encounter complex, L3Mo,CO, calculated to be without barrier and exothermic. While the overall reaction to form the C,MoL3 and O,MoL3 products was calculated to be energetically favorable, both the encounter complex and intermediate dimer, L3Mo,CO,MoL3, were found to be lower in energy than the products, with the final C,O cleavage step calculated to be endothermic by 169 kJ,mol,1 and 163 kJ,mol,1 for L = NH2 and N(tBu)Ar, respectively. The unfavorable CO cleavage step can be attributed to the fact that Mo does not possess the optimum d-electron configuration to sufficiently stabilise the carbide and oxide products relative to the CO-bridged intermediate dimer.(© Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2007) [source] Hydrolysis of the amyloid ,-peptide (A,) 1,40 between Asp23,Val24 produces non-aggregating fragments.JOURNAL OF MASS SPECTROMETRY (INCORP BIOLOGICAL MASS SPECTROMETRY), Issue 2 2005An electrospray mass spectrometric study Abstract The aggregation of full-length (residues 1,40) amyloid ,-peptide (A,) and fragments corresponding to residues 1,23 and 24,40 was studied by electrospray mass spectrometry, using gramicidin as a non-aggregating reference. Following a lag period, A,(1,40) at 140 µM concentration aggregates with apparent first-order kinetics. Under acidic conditions A,(1,40) undergoes spontaneous cleavage between Asp23,Val24 and to a lesser extent also at two other Asp,X motifs. Incubation in acidic H218O showed incorporation of 18O in fragment A,(1,23), confirming that the Asp23,Val24 peptide bond had been hydrolyzed. Incubation of synthetic A,(1,23) and A,(24,40) peptides with A,(1,40) showed that A,(24,40) remained in solution for several months, that A,(1,23) partly disappeared from solution, whereas A,(1,40) completely disappeared. Further, treatment of sedimentable aggregates formed after co-incubation of the three peptides with hexafluoro-2-propanol or formic acid recovered the intensity of A,(1,40). These data support previous studies showing that the region of A, encompassing residues 16,24 is necessary for aggregation into amyloid fibrils. Copyright © 2005 John Wiley & Sons, Ltd. [source] Construction of the Active Site of Metalloenzyme on Au NC MicellesCHINESE JOURNAL OF CHEMISTRY, Issue 7 2009Zhiming ZHANG Abstract For developing an efficient nanoenzyme system with self-assembly strategy, gold nanocrystal micelles (Au NC micelles) with inserted catalytic Zn(II) centers were constructed by self-assembly of a catalytic ligand [N,N -bis(2-aminoethyl)- N, -dodecylethylenediamine] Zn(II) complexes (Zn(II)L) on the surface of Au NC via hydrophobic interaction. The functionalized Au NC micelles acted as an excellent nanoenzyme model for imitating ribonuclease. The catalytic capability of the Au NC micelles was evaluated by accelerating the cleavage of 2-hydroxypropyl p -nitrophenyl phosphate (HPNP). These functionalized Au NC micelles exhibited considerable ribonuclease-like activities by a factor of 4.9×104 (kcat/kuncat) for the cleavage of HPNP in comparison to the spontaneous cleavage of HPNP at 37°C. The catalytic capability of the functionalized Au NC micelles can be considerably compared to other models reported previously as nanoenzymes under the comparable conditions. [source] | ||||||||||