The Application of 199 Hg NMR and 199m Hg Perturbed Angular Correlation (PAC) Spectroscopy to Define the Biological Chemistry of Hg II : A Case Study with Designed Two- and Three-Stranded Coiled Coils

The use of de novo designed peptides is a powerful strategy to elucidate Hg II –protein interactions and to gain insight into the chemistry of Hg II in biological systems. Cysteine derivatives of the designed Α-helical peptides of the TRI family [Ac-G-(L a K b A c L d E e E f K g ) 4 -G-NH 2 ] bind Hg II at high pH values and at peptide/Hg II ratios of 3:1 with an unusual trigonal thiolate coordination mode. The resulting Hg II complexes are good water-soluble models for Hg II binding to the protein MerR. We have carried out a parallel study using 199 Hg NMR and 199m Hg perturbed angular correlation (PAC) spectroscopy to characterize the distinct species that are generated under different pH conditions and peptide TRI 14L9C/Hg II ratios. These studies prove for the first time the formation of [Hg{(TRI 14L9C) 2 -(TRI 14L9C[bond]H)}], a dithiolate–Hg II complex in the hydrophobic interior of the three-stranded coiled coil (TRI 14L9C) 3 . 199 Hg NMR and 199m Hg PAC data demonstrate that this dithiolate–Hg II complex is different from the dithiolate [Hg(TRI 14L9C) 2 ], and that the presence of third Α-helix, containing a protonated cysteine, breaks the symmetry of the coordination environment present in the complex [Hg(TRI 14L9C) 2 ]. As the pH is raised, the deprotonation of this third cysteine generates the trigonal thiolate–Hg II complex Hg(TRI 14L9C) 3 − on a timescale that is slower than the NMR timescale (0.01–10 14ms). The formation of the species [Hg{(TRI 14L9C) 2 (TRI 14L9C[bond]H)}] is the result of a compromise between the high affinity of Hg II to form dithiolate complexes and the preference of the peptide to form a three-stranded coiled coil.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/57382/1/9178_ftp.pd

The Correlation of 113 Cd NMR and 111m Cd PAC Spectroscopies Provides a Powerful Approach for the Characterization of the Structure of Cd II -Substituted Zn II Proteins PAC Spectroscopy = perturbed angular correlation spectroscopy

The powerful combination of 113 Cd 14NMR and 111m Cd PAC (perturbed angular correlation) spectroscopies has been critical to determine the coordination geometry of Cd II bound to thiolate-rich centers. We have obtained important linear correlations between 113 Cd 14NMR and 111m Cd PAC spectroscopic data and the acid/base properties of the metal binding site that illustrate the presence of a dynamic model for metal binding (see figure). These unique results can give new insight into Cd II -substituted Zn II proteins. Cd II has been used as a probe of zinc metalloenzymes and proteins because of the spectroscopic silence of Zn II . One of the most commonly used spectroscopic techniques is 113 Cd 14NMR; however, in recent years 111m Cd Perturbed Angular Correlation spectroscopy ( 111m Cd PAC) has also been shown to provide useful structural, speciation and dynamics information on Cd II complexes and biomolecules. In this article, we show how the joint use of 113 Cd 14NMR and 111m Cd PAC spectroscopies can provide detailed information about the Cd II environment in thiolate-rich proteins. Specifically we show that the 113 Cd 14NMR chemical shifts observed for Cd II in the designed TRI series (TRI=Ac-G(LKALEEK) 4 G-NH 2 ) of peptides vary depending on the proportion of trigonal planar CdS 3 and pseudotetrahedral CdS 3 O species present in the equilibrium mixture. PAC spectra are able to quantify these mixtures. When one compares the chemical shift range for these peptides (from Δ =570 to 700 14ppm), it is observed that CdS 3 species have Δ 675–700 14ppm, CdS 3 O complexes fall in the range Δ 570–600 14ppm and mixtures of these forms fall linearly between these extremes. If one then determines the p K a2 values for Cd II complexation [p K a2 is for the reaction Cd[(peptide−H) 2 (peptide)] + →Cd(peptide) 3 − + 2H + ] and compares these to the observed chemical shift for the Cd(peptide) 3 − complexes, one finds that there is also a direct linear correlation. Thus, by determining the chemical shift value of these species, one can directly assess the metal-binding affinity of the construct. This illustrates how proteins may be able to fine tune metal-binding affinity by destabilizing one metallospecies with respect to another. More important, these studies demonstrate that one may have a broad 113 Cd 14NMR chemical shift range for a chemical species (e.g., CdS 3 O) which is not necessarily a reflection of the structural diversity within such a four-coordinate species, but rather a consequence of a fast exchange equilibrium between two related species (e.g., CdS 3 O and CdS 3 ). This could lead to reinterpretation of the assignments of cadmium–protein complexes and may impact the application of Cd II as a probe of Zn II sites in biology.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/62050/1/3761_ftp.pd

Consideraciones de las dinámicas y procesos de cambio de población y del ambiente para el desarrollo territorial con un sistema alimentario y nutricional sostenible Centroamericano

Los análisis de dinámicas y procesos de cambio territorial nos per-miten interpretar y dar sentido a la articulación entre los diferentes elementos constitutivos de los ámbitos del desarrollo y cómo estos se han generado, integrando en los procesos una visión dinámica y prospectiva muy pertinente para abordajes integrales, incluyentes y sostenibles del desarrollo territorial. En el presente escrito se abor-dan los ámbitos de población y del ambiente desde una perspectiva regiona