Fluorescence and FT-IR spectroscopic studies of Suwannee River fulvic acid complexation with aluminum, terbium and calcium.

In this study fluorescence emission and IR spectroscopy have been used to investigate the interaction of the class A (oxygen seeking "hard acid") metal Al(3+), with Suwannee River fulvic acid. Addition of Al(3+) ion results in a significant enhancement in fulvic acid fluorescence emission (at lambda(em)=424 nm) and significant red shift of the excitation wavelength (from lambda(ex)=324 nm to lambda(ex)=344 nm) at low pH values (pH approximately 4.0-5.0). At pH 4.0 (0.1 M ionic strength), where the predominant aluminum ion species is the "free" (aquo) ion, the fulvic acid fluorescence reaches 142% of the value in the absence of added metal ion. Analysis of the pH 4.0 and pH 5.0 fluorescence enhancement data with the nonlinear (single site) model of Ryan and Weber indicated binding constants in the range of 4.67.10(4)-2.87.10(6) M(-1) and concentrations of ligand sites in the range of 18.6.10(-6)-24.0.10(-6) M, both consistent with previous studies performed on both aquatic and soil fulvic acids. Companion fluorescence experiments performed on two other class A metal ions, Ca(2+) and Tb(3+), indicated no significant enhancement or quenching with Ca(2+) and only slight quenching with Tb(3+). Comparison of FT-IR spectra collected on fulvic acid alone and fulvic acid in the presence of the three class A metals (Al(3+), Ca(2+) and Tb(3+)) provides strong evidence for the involvement of carboxyl carbonyl functions in the binding of all three metal ions, which is not unexpected. The spectra also reveal, however, a very pronounced difference in the 4000-2000 cm(-1) IR spectral region between the Al(3+) spectrum and the Ca(2+) and Tb(3+) spectra. The -OH stretch spectral region in the Al(3+) spectrum has a major component shifted to higher energy (compared to fulvic acid alone or to fulvic acid in the presence of Ca(2+) or Tb(3+)). Even more striking is the emergence of a pronounced IR band at 2407 cm(-1), which is present only in the Al(3+) spectrum. The results of fluorescence and IR experiments with the model compounds salicylic acid and phthalic acid further confirm that both salicylic acid-like sites and phthalic acid-like sites are likely complexation sites for Al(3+) in fulvic acid and are major contributors to the observed spectroscopic changes associated with Al(3+) ion complexation. From a comparison of both the fluorescence and IR spectral results for all three class A metals, differing most strongly in the value of their ionic index, it seems clear that major sources of the deviation in spectral properties between Al(3+) and Ca(2+)/Tb(3+) is the unusually high value of its charge density and relatively low propensity for involvement in covalent bonding interactions (very high ionic index and relatively low covalent index in the Nieboer and Richardson classification of environmental metals), as well as affinity for certain functional groups.

Molecular dynamics study of Ca(2+) binding loop variants of parvalbumin with modifications at the "gateway" position.

The helix-loop-helix (i.e. EF-hand) Ca(2+) ion binding motif is characteristic of a large family of high-affinity Ca(2+) ion binding proteins, including the parvalbumins and calmodulins. In this paper we describe a set of molecular dynamics computations on the major parvalbumin from the silver hake (SHPV-B). In all variants examined, both whole protein and fragments thereof, the ninth loop residue in the Ca(2+) binding coordination site in the CD helix-loop-helix region (the so-called "gateway" residue) has been mutated. The three gateway mutations examined are arginine, which has never been found at the gateway position of any EF-hand protein, cysteine, which is the residue observed least in natural EF-hand sites, and serine, which is the most common (by far) non-acidic residue substitution at this position in EF-hand proteins in general, but never in parvalbumins. Results of the molecular dynamics simulations indicate that all three modifications are disruptive to the integrity of the mutated Ca(2+) binding site in the whole parvalbumin protein. In contrast, only the arginine and cysteine mutations are disruptive to the integrity of the mutated Ca(2+) binding site in the CD fragment of the parvalbumin protein. Surprisingly, the serine variant of the CD helix-loop-helix fragment exhibited remarkable stability during the entire molecular dynamics simulation, with retention of the Ca(2+) binding site. These results indicate that there are no inherent problems (for Ca(2+) ion binding) associated with the sequence of the CD helix-loop-helix fragment that precludes the incorporation of serine at the gateway position. Since the CD site is totally disrupted in the whole protein serine variant, this indicates that the Ca(2+) ion binding deficiencies are most likely related to the unique interaction that exists between the paired EF-hands in the whole protein. Our theoretical results correlate well with previous studies on engineered EF-hand proteins and with all of our experimental evidence on the silver hake parvalbumin.

Spatial working memory score of humans in a large radial maze, similar to published score of rats, implies capacity close to the magical number 7 +/- 22.

To compare the working memory (WM) capacity of humans to rats, we tested humans with a 17-arm radial maze and, in a follow up experiment, with a 13-arm radial maze. Both mazes were 15.2 meters in diameter, painted on a grassy field. In one version of the 13-arm experiment, we required a concurrent nonsense vocalization to impede subjects' use of language to remember locations. Subjects were instructed to choose arms of the radial maze unsystematically--as rats generally appear to do--and to visit the end of each arm only once. In additional procedures, we tested working memory capacity in a verbal task that is more analogous to the radial maze than is the typical ordered recall test. Subjects were asked to try to recite a sequence of 17 numbers (i.e., 18 through 34) or letters (A through Q) in unsystematic order, with no repeats. In another experiment subjects recited 13 numbers (14-26) or letters (A-M). In all tests, subjects were allowed only as many responses as there were distinct items (17 or 13, respectively). Average correct-response (nonrepeat) scores were 14.4 for the 17-arm maze and 14.1 for both of the verbal 17-item tests; these scores are close to the reported score for rats in a 17-arm radial maze. Average scores were between 10.8 and 11.4 in all of the 13-item maze and recitation tasks.(ABSTRACT TRUNCATED AT 250 WORDS)