Nathan Yee

A chemical equilibrium model for metal adsorption onto bacterial surfaces

Abstract This study quantifies metal adsorption onto cell wall surfaces of Bacillus subtilis by applying equilibrium thermodynamics to the specific chemical reactions that occur at the water-bacteria interface. We use acid/base titrations to determine deprotonation constants for the important surface functional groups, and we perform metal-bacteria adsorption experiments, using Cd, Cu, Pb, and Al, to yield site-specific stability constants for the important metal-bacteria surface complexes. The acid/base properties of the cell wall of B. subtilis can best be characterized by invoking three distinct types of surface organic acid functional groups, with pK a values of 4.82 ± 0.14, 6.9 ± 0.5, and 9.4 ± 0.6. These functional groups likely correspond to carboxyl, phosphate, and hydroxyl sites, respectively, that are displayed on the cell wall surface. The results of the metal adsorption experiments indicate that both the carboxyl sites and the phosphate sites contribute to metal uptake. The values of the log stability constants for metal-carboxyl surface complexes range from 3.4 for Cd, 4.2 for Pb, 4.3 for Cu, to 5.0 for Al. These results suggest that the stabilities of the metal-surface complexes are high enough for metal-bacterial interactions to affect metal mobilities in many aqueous systems, and this approach enables quantitative assessment of the effects of bacteria on metal mobilities.

Cd adsorption onto bacterial surfaces: A universal adsorption edge?

In this study, we measure the thermodynamic stability constants for proton and Cd binding onto the Gram-negative bacteria Escherichia coli and Pseudomonas aeruginosa, and the Gram-positive bacteria Bacillus megaturium, Streptococcus faecalis, Staphylococcus aureus, Sporosarcina ureae, and Bacillus cereus. Potentiometric titrations and Cd-bacteria adsorption experiments yield average values for the carboxyl site pKa, site concentration, and log stability constant for the bacterial surface Cd-carboxyl complex of 5.0, 2.0 × 10−3 mol/g and 4.0 respectively. Our results indicate that a wide range of bacterial species exhibit nearly identical Cd adsorption behavior as a function of pH. We propose that metal-bacteria adsorption is not dependent on the bacterial species involved, and we develop a generalized adsorption model which may greatly simplify the task of quantifying the effects of bacterial adsorption on dissolved mass transport in realistic geologic systems.

X-ray absorption fine-structure determination of pH-dependent U-bacterial cell wall interactions.

X-ray absorption fine structure (XAFS) measurements was used at the U L3-edge to directly determine the pH dependence of the cell wall functional groups responsible for the absorption of aqueous UO22+ to Bacillus subtilis from pH 1.67 to 4.80. Surface complexation modeling can be used to predict metal distributions in water–rock systems, and it has been used to quantify bacterial adsorption of metal cations. However, successful application of these models requires a detailed knowledge not only of the type of bacterial surface site involved in metal adsorption/desorption, but also of the binding geometry. Previous acid-base titrations of B. subtilis cells suggested that three surface functional group types are important on the cell wall; these groups have been postulated to correspond to carboxyl, phosphoryl, and hydroxyl sites. When the U(VI) adsorption to B. subtilis is measured, observed is a significant pH-independent absorption at low pH values (<3.0), ascribed to an interaction between the uranyl cation and a neutrally charged phosphoryl group on the cell wall. The present study provides independent quantitative constraints on the types of sites involved in uranyl binding to B. subtilis from pH 1.67 to 4.80. The XAFS results indicate that at extremely low pH (pH 1.67) UO22+ binds exclusively to phosphoryl functional groups on the cell wall, with an average distance between the U atom and the P atom of 3.64 ± 0.01 A. This U-P distance indicates an inner-sphere complex with an oxygen atom shared between the UO22+ and the phosphoryl ligand. The P signal at extremely low pH value is consistent with the UO22+ binding to a protonated phosphoryl group, as previously ascribed. With increasing pH (3.22 and 4.80), UO22+ binds increasingly to bacterial surface carboxyl functional groups, with an average distance between the U atom and the C atom of 2.89 ± 0.02 A. This U-C distance indicates an inner-sphere complex with two oxygen atoms shared between the UO22+ and the carboxyl ligand. The results of this XAFS study confirm the uranyl-bacterial surface speciation model.

Experimental study of the pH, ionic strength, and reversibility behavior of bacteria–mineral adsorption

In this study, we investigate the adsorption of Bacillus subtilis onto the surfaces of two minerals, corundum and quartz, as a function of time, pH, ionic strength, and bacteria:mineral mass ratio. Experimental results indicate that the adsorption of bacteria onto a mineral surface is a completely reversible process with equilibrium being reached in less than 1 h. Our data also indicate that B. subtilis displays a higher affinity for corundum surfaces than for quartz surfaces, and that the extent of bacteria adsorption onto corundum increases with decreasing pH, with increasing bacteria:mineral mass ratio, and with decreasing ionic strength. The adsorption behavior is governed by the chemical speciation of the bacterial and mineral surfaces. We describe the experimental results with a chemical equilibrium model. The model accounts for hydrophobic and electrostatic interactions that occur between the bacteria and mineral surfaces, and can account for the effects of solution chemistry as well as surface speciation on the extent of adsorption. These results are the first to integrate the effects of pH, ionic strength, and bacteria:mineral ratio in a quantitative model. Such an approach enables bacteria-mineral adsorption reactions to be incorporated into more standard water-rock speciation models, providing a better understanding of mass transport in both natural and bio-engineered bacteria-bearing geochemical systems. Copyright

A comparison of the thermodynamics of metal adsorption onto two common bacteria

The cell walls of bacteria are known to adsorb a variety of metals, and thus they may control metal mobilities in many low-temperature aqueous systems. In order to quantify metal adsorption onto bacterial surfaces, recent studies have applied equilibrium thermodynamics to the specific chemical and electrostatic interactions occurring at the solution–cell wall interface. However, to date, few studies have used this approach to compare the surface properties and metal affinities of different species of bacteria. In this study, we use acid–base titrations to determine the concentrations and deprotonation constants of specific surface functional groups on Bacillus licheniformis. The cell wall displays carboxyl, phosphate and hydroxyl surface functional groups, with pKa values and 1s errors of 5.2±0.3, 7.5±0.4 and 10.2±0.5, respectively. We perform metal–B. licheniformis adsorption experiments using Cd, Pb, Cu and Al. The average log K values for the Cd-, Pb-, Cu- and Al–carboxyl stability constants, with 1s errors, are 3.9±0.5, 4.6±0.3, 4.9±0.4 and 5.8±0.3, respectively. Finally, we compare the surface characteristics and metal affinities of B. licheniformis to those of Bacillus subtilis, as determined by Fein et al. [Fein, J.B., Daughney, C.J., Yee, N., Davis, T., 1997. A chemical equilibrium model of metal adsorption onto bacterial surfaces. Geochim. Cosmochim. Acta 61, 3319–3328]. Our investigations indicate that these two species of bacteria have different relative and absolute concentrations of surface sites and slightly different deprotonation and metal adsorption stability constants. We relate these variations in surface properties to variations in metal affinity in order to predict metal mobilities in complex, natural systems.

The effect of cyanobacteria on silica precipitation at neutral pH: implications for bacterial silicification in geothermal hot springs

In this study, we performed silica precipitation experiments with the cyanobacteria Calothrix sp. to investigate the mechanisms of silica biomineralization. Batch silica precipitation experiments were conducted at neutral pH as a function of time, Si saturation states, temperature and ferrihydrite concentrations. The experimental results show that in solutions undersaturated with respect to amorphous silica, the interaction between Si and cell surface functional groups is weak and minimal Si sorption onto cyanobacterial surfaces occurs. In solutions at high Si supersaturation states, abiotic Si polymerization is spontaneous, and at the time scales of our experiments (1–50 h) the presence of cyanobacteria had a negligible effect on silica precipitation kinetics. At lower supersaturation states, Si polymerization is slow and the presence of cyanobacteria do not promote Si–solid phase nucleation. In contrast, experiments conducted with ferrihydrite-coated cyanobacteria significantly increase the rate of Si removal, and the extent to which Si is removed increases as a function of ferrihydrite concentration. Experiments conducted with inorganic ferrihydrite colloids (without cyanobacteria) removes similar amounts of Si, suggesting that microbial surfaces play a limited role in the silica precipitation process. Therefore, in supersaturated hydrothermal waters, silica precipitation is largely nonbiogenic and cyanobacterial surfaces have a negligible effect on silica nucleation. D 2003 Elsevier Science B.V. All rights reserved.

Molecular characterization of cyanobacterial silicification using synchrotron infrared micro-spectroscopy

Synchrotron-based Fourier-transform infrared (SR-FTIR) micro-spectroscopy was used to determine the concentration-dependent response of the organic structure of live cyanobacterial cells to silicification. Mid-infrared (4000–600 cm−1) measurements carried out on single filaments and sheaths of the cyanobacteria Calothrix sp. (strain KC97) were used to monitor the interaction between a polymerizing silica solution and the organic functional groups of the cells during progressive silicification. Spectra of whole-cells and sheaths were analyzed and the spectral features were assigned to specific functional groups related to the cell: lipids (-CH2 and -CH3; at 2870–2960 cm−1), fatty acids (>C=O at 1740 cm−1), proteins (amides I and II at 1650 and 1540 cm−1), nucleic acids (>P=O 1240 cm−1), carboxylic acids (C-O at 1392 cm−1), and polysaccharides (C-O between 1165 and 1030 cm−1). These vibrations and the characteristic vibrations for silica (Si-O between 1190 and 1060 cm−1; to some extent overlapping with the C-O frequencies of polysaccharides and Si-O at 800 cm−1) were used to follow the progress of silicification. Relative to unsilicified samples, the intensity of the combined C-O/Si-O vibration band increased considerably over the course of the silicification (whole-cells by > 90% and sheath by ∼75%). This increase is a consequence of (1) extensive growth of the sheath in response to the silicification, and (2) the formation of thin amorphous silica layers on the sheath. The formation of a silica specific band (∼800 cm−1) indicates, however, that the precipitation of amorphous silica is controlled by the dehydroxylation of abiotically formed silanol groups.

The dynamics of cyanobacterial silicification: an infrared micro-spectroscopic investigation

Abstract The dynamics of cyanobacterial silicification was investigated using synchrotron-based Fourier transform infrared micro-spectroscopy. The changes in exo-polymeric polysaccharide and silica vibrational characteristics of individual Calothrix sp. filaments was determined over time in a series of microcosms in which the microbially sorbed silica or silica and iron load was increased sequentially. The changes in intensity and integrated area of specific infrared spectral features were used to develop an empirical quantitative dynamic model and to derive silica load-dependent parameters for each quasi-equilibrium stage in the biomineralization process. The degree of change in spectral features was derived from the increase in integrated area of the combined silica/polysaccharide region (Si-O/C-O, at 1150–950 cm−1) and the Si-O band at 800 cm−1, the latter representing specific silica bonds corresponding to hydrated amorphous SiO4 tetrahedra. From the degree of change, a two-phase model with concurrent change in process was derived. In the first phase, a biologically controlled increase in thickness of the exo-polymeric polysaccharide sheath around the cell was observed. In phase two, a transition to an inorganically controlled accumulation of silica on the surface of the cyanobacterial cells was derived from the change in integrated area for the mixed Si-O/C-O spectral region. This second process is further corroborated by the synchronous formation of non-microbially associated inorganic SiO4 units indicated by the growth of the singular Si-O band at 800 cm−1. During silicification, silica accumulates (1) independently of the growth of the sheath polysaccharides and (2) via an increase in chain lengths of the silica polymers by expelling water from the siloxane bonds. IR evidence suggest that an inorganic, apparently surface catalyzed process, which leads to the accumulation of silica nanospheres on the cyanobacterial surfaces governs this second stage. In experiments where iron was present, the silicification followed similar pathways, but at low silica loads, the iron bound to the cell surfaces slightly enhanced the reaction dynamics.

The rate of ferrihydrite transformation to goethite via the Fe(II) pathway

Abstract In this study, we quantified the rate of ferrihydrite conversion to goethite via the Fe(II) pathway using synchrotron radiation-based energy dispersive X-ray diffraction (ED-XRD). Ferrihydrite transformation experiments were conducted in oxygen-free solutions at neutral pH with synthetic 2-line ferrihydrite reacting with 100 mM Fe(II). The kinetics of goethite crystallization was measured in situ at temperatures ranging from 21 to 90 °C. The results showed that in the presence of ferrous iron, the transformation of poorly ordered ferrihydrite into crystalline goethite is rapid and highly dependent on temperature. The time-resolved peak area data fitted using a Johnson-Mehl-Avrami-Kolmogorov (JMAK) kinetic model yielded rate constants of 4.0 x 10-5, 1.3 x 10-4, 3.3 x 10-4, 2.27 x 10-3, and 3.14 x 10-3 1/s at reaction temperatures of 21, 45, 60, 85, and 90 °C respectively. The activation energy for the transformation was determined to be 56 ± 4 kJ/mol. Comparison with the activation energy predicted for the phase conversion in the absence of ferrous iron indicates that Fe(II) acts as a catalyst that decreases the activation energy barrier by approximately 38 kJ/mol. The kinetic parameters derived from the experimental data suggest that goethite crystallization is controlled by a 1-D phase boundary growth mechanism with a constant nucleation rate occurring during the reaction

Se(VI) Reduction and the Precipitation of Se(0) by the Facultative Bacterium Enterobacter cloacae SLD1a-1 Are Regulated by FNR

ABSTRACT The fate of selenium in the environment is controlled, in part, by microbial selenium oxyanion reduction and Se(0) precipitation. In this study, we identified a genetic regulator that controls selenate reductase activity in the Se-reducing bacterium Enterobacter cloacae SLD1a-1. Heterologous expression of the global anaerobic regulatory gene fnr (fumarate nitrate reduction regulator) from E. cloacae in the non-Se-reducing strain Escherichia coli S17-1 activated the ability to reduce Se(VI) and precipitate insoluble Se(0) particles. Se(VI) reduction by E. coli S17-1 containing the fnr gene occurred at rates similar to those for E. cloacae, with first-order reaction constants of k = 2.07 × 10−2 h−1 and k = 3.36 × 10−2 h−1, respectively, and produced elemental selenium particles with identical morphologies and short-range atomic orders. Mutation of the fnr gene in E. cloacae SLD1a-1 resulted in derivative strains that were deficient in selenate reductase activity and unable to precipitate elemental selenium. Complementation by the wild-type fnr sequence restored the ability of mutant strains to reduce Se(VI). Our findings suggest that Se(VI) reduction and the precipitation of Se(0) by facultative anaerobes are regulated by oxygen-sensing transcription factors and occur under suboxic conditions.