H3PO4 activated Typha angustifolia and Salix matsudana carbons provide good adsorption/desorption characteristics for Cd and Pb.

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Unfolding and aggregation are basic problems in protein science with serious biotechnological and medical implications. Probing the sequential events occurring during the unfolding and aggregation process and the relationship between unfolding and aggregation is of particular interest. In this study, two-dimensional infrared (2D IR) correlation spectroscopy was used to study the sequential events and starting temperature dependence of Myoglobin (Mb) thermal transitions. Though a two-state model could be obtained from traditional 1D IR spectra, subtle noncooperative conformational changes were observed at low temperatures. Formation of aggregation was observed at a temperature (50-58 degrees C) that protein was dominated by native structures and accompanied with unfolding of native helical structures when a traditional thermal denaturation condition was used. The time course NMR study of Mb incubated at 55 degrees C for 45 h confirmed that an irreversible aggregation process existed. Aggregation was also observed before fully unfolding of the Mb native structure when a relative high starting temperature was used. These findings demonstrated that 2D IR correlation spectroscopy is a powerful tool to study protein aggregation and the protein aggregation process observed depends on the different environmental conditions used.

The kinetics of the reversible thermal unfolding, irreversible thermal unfolding, and reductive unfolding processes of bovine pancreatic ribonuclease A (RNase A) were investigated in NaCl/P i solutions. Image parameters including Shannon entropy, Hamming distance, mutual information and correlation coefficient were used in the analysis of the CD and 1D NMR spectra. The irreversible thermal unfolding transition of RNase A was not a cooperative process, pretransitional structure changes occur before the main thermal denaturation. Different dithiothreitol (dithiothreitol red ) concentration dependencies were observed between 303 and 313 K during denaturation induced by a small amount of reductive reagent. The protein selectively follows a major unfolding kinetics pathway with the selectivity can be altered by temperature and reductive reagent concentration. Two possible explanations of the selectivity mechanism were discussed.Keywords: image analysis; proton nuclear magnetic resonance; reductive unfolding; thermal unfolding; unfolding kinetics.Dynamic analysis of the unfolding and refolding pathways and identification of the specific conformational changes which form the individual intermediates involved in the rate-limited pathway(s) can distinguish one pathway from another and play fundamental importance for protein folding [1,2]. It is usually of considerable interest to estimate the conformational changes of both the whole protein tertiary structure and of specific sites observed by spectroscopic techniques in different redox systems and solvent conditions. Protein unfolding is highly pertinent to protein folding [3] and is more controllable for more comprehensive study by slowing the unfolding process carried out at physiological pH and temperature [4]. Such studies in turn provide new insights into the functional properties and mechanisms of proteins, which will lead to a more detailed and more complete description of biological functions [5].Bovine pancreatic ribonuclease A (RNase A; EC 3.1.27.5) contains 124 residues with four native disulfide bonds (Cys26-Cys84, Cys40-Cys95, Cys58-Cys110, and Cys65-Cys72). RNase A has played a crucial role as a model system in studies of protein structure, folding, stability, and chemistry [6]. It folds and unfolds through multiple pathways, with the rate-limiting steps in the wellpopulated pathways involving the formation of distinct transition intermediates [1][2][3]7,8]. The complexity of the multiple pathways means that different mechanisms may occur with different types of redox systems and different solvent conditions [9,10]. Thus a comprehensive investigation under different conditions using different methods is essential to elucidate the protein folding and unfolding processes.Much effort has been devoted in recent years to understanding the mechanism and the main factors that control protein folding, and to developing approaches that allow researchers to investigate the multifarious aspects of protein folding and unfolding. While the determination of the protein s...

The dynamics of the unfolding process of bovine pancreatic ribonuclease A (RNase A) unfolded by dithiothreitol (DTT) at a low concentration of 1:30 were investigated in alkaline phosphate-buffered saline solutions at 303K and 313K by using proton nuclear magnetic resonance ( 1 H NMR) spectra. Three NMR spectral parameters including Shannon entropy, mutual information, and correlation coefficient were introduced into the analysis. The results show that the unfolding process of RNase A was slowed to the order of many hours, and the kinetics of the unfolding pathway described by the three parameters is best fit by a model of two consecutive first-order reactions. Temperature greatly influences the rate constants of the unfolding kinetics with different temperature effects observed for the fast and the slow processes. The consistencies and the differences between the three sets of parameters show that they reflect the same relative denaturation pathway but different spectra windows of the unfolding process of RNase A. The results suggest that the unfolding process of RNase A induced by low concentrations of DTT is a two-phase pathway containing fast and slow first-order reactions.