Sulfur Allocation and Vanadium−Sulfate Interactions in Whole Blood Cells from the Tunicate Ascidia ceratodes, Investigated Using X-ray Absorption Spectroscopy

Abstract

Sulfur K-edge X-ray absorption spectroscopy (XAS) has been used to investigate the distribution of sulfur types in two whole blood cell samples, in selected subcellular blood fractions, and in cell-free plasma from the tunicate Ascidia ceratodes. Whole blood cells are rich in sulfate, aliphatic sulfonate, and low-valent sulfur. The sulfur K-edge XAS spectrum of washed blood cell membranes revealed traces of sulfate and low-valent sulfur, but no sulfate ester or sulfonate. Sulfonate is thus exclusively cytosolic. Cell-free blood plasma contains primarily sulfate sulfur. Gaussian fitting and sulfate quantitation for two whole blood cell samples, each representing dozens of individuals from separate collections taken per annum, yielded average sulfur type concentrations for the two populations: (first year, second year): [sulfate], 110 mM, 150 mM; [sulfonate], 99 mM, 70 mM, and; [low-valent sulfur], 41 mM, 220 mM. On titration of sulfate with acid or V(III) in aqueous solution, the strong 2482.4 eV 1s → (valence t 2 ) sulfur K-edge XAS transition of tetrahedral SO4 2- splits into 1s → a1 and 1s → e transitions, because HSO4 - and VSO4 + are of C 3 v symmetry. Gaussian fits and appropriate comparisons allow the following assignments: (compound/complex, 1s → a1 (eV), 1s → e (eV)): myo-inositol hexasulfate, 2480.8, 2482.8; HSO4 -, 2481.4, 2482.7; VSO4 +, 2481.2, 2482.9. The energy separating the a1 and e states of complexed sulfate appears to be solvation dependent. From these studies is derived an explicit inorganic spectrochemical model for biological V(III) and sulfate. The average endogenous equilomer concentrations of sulfate complexed with V(III) and/or H+ within the two blood cell samples are calculated from this model. The results provide a natural explanation for the observed biological broadening of A. ceratodes blood cell sulfur K-edge XAS spectra.

The uptake and fate of vanadyl ion in ascidian blood cells and a detailed hypothesis for the mechanism and location of biological vanadium reduction. A visible and X-ray absorption spectroscopic study

“…N(2-hydroxyethyl)-N,N′,N′-triacetic acid) 5 ], i.e., the principle sulfate complex ion of V(III), decreased even more than would be expected by the absolute increase in intracellular vanadium due to endocytosed vanadyl ion. If the aqua and sulfated forms of V(III) reside in the same vacuole, as is virtually certain [9,12,15,38,42,55], then this reciprocal response is consistent with an increase in acidity following uptake of vanadyl ion. Such an increase in acidity could be accomplished either by influx of [H + ] to the vacuole, or efflux of [SO 4 2− ] from the vacuole.…”

“…Each final spectrum represents the average of 16 to 20 individual scans. The vanadium K-edge XAS spectra of all the solid and solution models used herein have been reported previously [15,22,26,31,38].…”

“…However, elements of glutathione metabolism have been found in ascidians [75,76], and a glutathione reductant is not inconsistent with the copious reduced sulfur detected in whole blood cells using sulfur Kedge XAS [12,15,38]. The successful chemical production of 7-coordinate V(III) from complexed VO 2+ by ascorbate or cysteine methyl ester, but not by NADPH [50], implies that a direct metal-reductant interaction is necessary.…”

“…Each final spectrum represents the average of 16 to 20 individual scans. The vanadium K-edge XAS spectra of all the solid and solution models used herein have been reported previously [15,22,26,31,38].Fits to the vanadium K-edge spectra were carried out using the program DATFIT4, which is part of the EXAFSPAK software package mentioned above. The blood cell vanadium K-edge spectra were first further processed within the program Kaleidagraph (Synergy Software, Reading PA) to remove extraneous curvature before carrying out the fits.…”

Vanadium haloperoxidases: From the discovery 30 years ago to X-ray crystallographic and V K-edge absorption spectroscopic studies

“…In subsequent studies, the V XANES features were used to quantify the amounts of V(3+) ions associated with the biological catecholate-based chelator tunichrome, and water and sulphate, as well as mixtures of the latter in various species [114][115][116][117][118], and a detailed mechanism for the reduction of vanadate to V(3+) was proposed [119]. In this particular context, the role of sulphur in the vanadium metabolism was also investigated with sulphur K edge XANES [120][121][122]. Micro-…”