Radiation maps of ocean sediment from the Castle Bravo crater.

On March 1, 1954, the United States conducted its largest thermonuclear weapon test in Bikini Atoll in the Marshall Islands; the detonation was code-named "Castle Bravo." Radioactive deposits in the ocean sediment at the bomb crater are widespread and high levels of contamination remain today. One hundred thirty cores were collected from the top 25 cm of surface sediment at ocean depths approaching 60 m over a ∼2-km2 area, allowing for a presentation of radiation maps of the Bravo crater site. Radiochemical analyses were performed on the following radionuclides: plutonium-(239,240), plutonium-238, americium-241, bismuth-207, and cesium-137. Large values of plutonium-(239,240), americium-241, and bismuth-207 are found. Comparisons are made to core sample results from other areas in the northern Marshall Islands.

Background gamma radiation and soil activity measurements in the northern Marshall Islands.

We report on measurements of external gamma radiation on 9 islands in 4 atolls in the northern Marshall Islands, all of which were affected by the US nuclear testing program from 1946 to 1958 (Enjebi, Ikuren, and Japtan in Enewetak Atoll; Bikini and Enyu in Bikini Atoll; Naen in Rongelap Atoll; and Aon, Elluk, and Utirik in Utirik Atoll). We also report americium-241, cesium-137, plutonium-238, and plutonium-239,240 activity concentrations in the soil samples for 11 islands in 4 northern atolls (Enewetak, Japtan, Medren, and Runit in Enewetak Atoll; Bikini and Enyu in Bikini Atoll; Naen and Rongelap in Rongelap Atoll; and Aon, Elluk, and Utirik in Utirik Atoll) and from Majuro Island, Majuro Atoll in the southern Marshall Islands. Our results show low external gamma radiation levels on some islands in the Enewetak Atoll and Utirik Atoll, and elevated levels on Enjebi Island in the Enewetak Atoll, on Bikini Atoll, and on Naen Island in the Rongelap Atoll. We perform ordinary kriging on external gamma radiation measurements to provide interpolated maps. We find that radionuclides are absent from all Majuro soil samples, and that they are present at highest activity concentrations in samples from Runit and Enjebi islands (Enewetak Atoll), Bikini Island (Bikini Atoll), and Naen Island (Rongelap Atoll). We contextualize all results by making comparisons between islands and to various standards, as well as to regions of the world affected by nuclear accidents. We also discuss implications for informed decision-making by the Marshallese and local atoll governments and their people on issues pertaining to island resettlement.

In situ measurement of cesium-137 contamination in fruits from the northern Marshall Islands.

Radioactive contamination of fruits in the northern Marshall Islands, resulting from the US nuclear weapons testing program in the 1940s and 1950s, is still a human health concern, in particular pertaining to island population resettlement and the economic benefit from farming. Over 200 fruits, primarily coconuts and pandanus, were collected on 11 islands from four atolls in the northern Marshall Islands in 2017. The energy spectra from nuclear gamma decays were measured on a research vessel for each fruit in situ. From these recordings, the level of cesium-137 (137Cs) contamination was determined for individual fruits. Comparisons of the results are made to past studies and international food safety standards. There is a broad distribution of values, ranging from below detectable radiation levels to relatively high levels; safety concerns are largest for Bikini Island. A noticeable fraction of fruits from Bikini have significantly higher levels of 137Cs contamination compared with those from all other measured islands.

Measurement of background gamma radiation in the northern Marshall Islands.

We report measurements of background gamma radiation levels on six islands in the northern Marshall Islands (Enewetak, Medren, and Runit onEnewetak Atoll; Bikini and Nam on Bikini Atoll; and Rongelap on Rongelap Atoll). Measurable excess radiation could be expected from the decay of (137)Cs produced by the US nuclear testing program there from 1946 to 1958. These recordings are of relevance to safety of human habitation and resettlement. We find low levels of gamma radiation for the settled island of Enewetak [mean = 7.6 millirem/year (mrem/y) = 0.076 millisievert/year (mSv/y)], larger levels of gamma radiation for the island of Rongelap (mean = 19.8 mrem/y = 0.198 mSv/y), and relatively high gamma radiation on the island of Bikini (mean = 184 mrem/y = 1.84 mSv/y). Distributions of gamma radiation levels are provided, and hot spots are discussed. We provide interpolated maps for four islands (Enewetak, Medren, Bikini, and Rongelap), and make comparisons to control measurements performed on the island of Majuro in the southern Marshall Islands, measurements made in Central Park in New York City, and the standard agreed upon by the United States and the Republic of the Marshall Islands (RMI) governments (100 mrem/y = 1 mSv/y). External gamma radiation levels on Bikini Island significantly exceed this standard (P = <<0.01), and external gamma radiation levels on the other islands are below the standard. To determine conclusively whether these islands are safe for habitation, radiation exposure through additional pathways such as food ingestion must be considered.

High-resolution analysis of Zn(2+) coordination in the alkaline phosphatase superfamily by EXAFS and x-ray crystallography.

Comparisons among evolutionarily related enzymes offer opportunities to reveal how structural differences produce different catalytic activities. Two structurally related enzymes, Escherichia coli alkaline phosphatase (AP) and Xanthomonas axonopodis nucleotide pyrophosphatase/phosphodiesterase (NPP), have nearly identical binuclear Zn(2+) catalytic centers but show tremendous differential specificity for hydrolysis of phosphate monoesters or phosphate diesters. To determine if there are differences in Zn(2+) coordination in the two enzymes that might contribute to catalytic specificity, we analyzed both x-ray absorption spectroscopic and x-ray crystallographic data. We report a 1.29-Å crystal structure of AP with bound phosphate, allowing evaluation of interactions at the AP metal site with high resolution. To make systematic comparisons between AP and NPP, we measured zinc extended x-ray absorption fine structure for AP and NPP in the free-enzyme forms, with AMP and inorganic phosphate ground-state analogs and with vanadate transition-state analogs. These studies yielded average zinc-ligand distances in AP and NPP free-enzyme forms and ground-state analog forms that were identical within error, suggesting little difference in metal ion coordination among these forms. Upon binding of vanadate to both enzymes, small increases in average metal-ligand distances were observed, consistent with an increased coordination number. Slightly longer increases were observed in NPP relative to AP, which could arise from subtle rearrangements of the active site or differences in the geometry of the bound vanadyl species. Overall, the results suggest that the binuclear Zn(2+) catalytic site remains very similar between AP and NPP during the course of a reaction cycle.

Probing the origin of the compromised catalysis of E. coli alkaline phosphatase in its promiscuous sulfatase reaction.

The catalytic promiscuity of E. coli alkaline phosphatase (AP) and many other enzymes provides a unique opportunity to dissect the origin of enzymatic rate enhancements via a comparative approach. Here, we use kinetic isotope effects (KIEs) to explore the origin of the 109-fold greater catalytic proficiency by AP for phosphate monoester hydrolysis relative to sulfate monoester hydrolysis. The primary 18O KIEs for the leaving group oxygen atoms in the AP-catalyzed hydrolysis of p-nitrophenyl phosphate (pNPP) and p-nitrophenylsulfate (pNPS) decrease relative to the values observed for nonenzymatic hydrolysis reactions. Prior linear free energy relationship results suggest that the transition states for AP-catalyzed reactions of phosphate and sulfate esters are "loose" and indistinguishable from that in solution, suggesting that the decreased primary KIEs do not reflect a change in the nature of the transition state but rather a strong interaction of the leaving group oxygen atom with an active site Zn2+ ion. Furthermore, the primary KIEs for the two reactions are identical within error, suggesting that the differential catalysis of these reactions cannot be attributed to differential stabilization of the leaving group. In contrast, AP perturbs the KIE for the nonbridging oxygen atoms in the reaction of pNPP but not pNPS, suggesting a differential interaction with the transferred group in the transition state. These and prior results are consistent with a strong electrostatic interaction between the active site bimetallo Zn2+ cluster and one of the nonbridging oxygen atoms on the transferred group. We suggest that the lower charge density of this oxygen atom on a transferred sulfuryl group accounts for a large fraction of the decreased stabilization of the transition state for its reaction relative to phosphoryl transfer.

Alkaline phosphatase catalysis is ultrasensitive to charge sequestered between the active site zinc ions.

Escherichia coli alkaline phosphatase (AP) is a prototypical bimetalloenzyme, facilitating catalysis of phosphate monoester hydrolysis with two Zn2+ metal ions that are only 4 A apart. In the reaction's transition state, one of the nonbridging oxygen atoms of the transferred group appears to interact directly with the Zn2+ ion metallocluster. To determine the importance and the energetic properties of this interaction, we systematically varied the charge on this oxygen atom, exploiting the ability of AP to catalyze reactions of different classes of substrates. We observed that the AP catalytic proficiency correlates very well (R2 = 0.98) with the charge on this oxygen atom, over 8 orders of magnitude of catalytic proficiency. The slope of this linear correlation (31 +/- 2 kcal/mol per unit charge) is extraordinarily steep, indicating that AP greatly discriminates between differentially charged substrates. We suggest that this discrimination arises via an electrostatic interaction with the bimetallocluster. The dependence of the AP catalytic proficiency on the nonbridging oxygen charge is much larger than charge perturbation effects observed previously for other proteins. We propose that AP uses folding energy to position the two Zn2+ metal ions in close proximity, thereby creating an active site with a high electrostatic potential that is extraordinarily sensitive to the charge that "solvates" the metallocluster. The sensitivity of enzyme energetics to systematic variation in electrostatic properties provides a powerful measure of the active site environment. Future work comparing the sensitivity of related enzymes that have been optimized to catalyze different reactions will help reveal how natural selection has tuned related active sites to favor different reactions.

Do electrostatic interactions with positively charged active site groups tighten the transition state for enzymatic phosphoryl transfer?

The effect of electrostatic interactions on the transition-state character for enzymatic phosphoryl transfer has been a subject of much debate. In this work, we investigate the transition state for alkaline phosphatase (AP) using linear free-energy relationships (LFERs). We determined k(cat)/K(M) for a series of aryl sulfate ester monoanions to obtain the Brønsted coefficient, beta(lg), and compared the value to that obtained previously for a series of aryl phosphorothioate ester dianion substrates. Despite the difference in substrate charge, the observed Brønsted coefficients for AP-catalyzed aryl sulfate and aryl phosphorothioate hydrolysis (-0.76 +/- 0.14 and -0.77 +/- 0.10, respectively) are strikingly similar, with steric effects being responsible for the uncertainties in these values. Aryl sulfates and aryl phosphates react via similar loose transition states in solution. These observations suggest an apparent equivalency of the transition states for phosphorothioate and sulfate hydrolysis reactions at the AP active site and, thus, negligible effects of active site electrostatic interactions on charge distribution in the transition state.

Environmental effects on phosphoryl group bonding probed by vibrational spectroscopy: implications for understanding phosphoryl transfer and enzymatic catalysis.

We have used vibrational spectroscopy to study bonding in monosubstituted dianionic phosphates, both to learn more about basic properties intrinsic to this important class of biological substrates and to assess the ability of vibrational spectroscopy to provide a "sensor" or probe of the local environment experienced by the phosphoryl group. We examined the bonding properties of the phosphoryl group via vibrational spectroscopy for a series of compounds in which the phosphoryl substituent was varied systematically and extensively. A broad linear correlation of the bridging P-O(R) bond length and the pK(a) of the substituent alcohol was observed. The results indicate that the P-O(R) bond changes by only approximately 0.04 A with alcohol substituents that vary in pK(a) by approximately 12 units, suggesting that phosphoryl group bonding responds in a subtle but regular manner to changes in the local environment. We also determined the effect on the phosphoryl bonding from changes in the solvent environment. Addition of dimethyl sulfoxide (DMSO) elongates the bridging bond, presumably as a result of lessened solvation to the nonbridging oxygens and conservation of bond order to phosphorus. Finally, we have addressed the relationship between ground-state bonding properties and reactivity, as changing the leaving group substituent and adding DMSO have large rate effects, and it was previously proposed that lengthening of the bond to be broken is the cause of the increased reactivity. The results herein suggest, however, that the change in the bridging bond energy is small compared to the changes in energy that accompany the observed reactivity differences. Further analysis indicates that electrostatic interactions can provide a common driving force underlying both bond lengthening and the observed rate increases. We suggest that ground-state distortions of substrates bound to enzymes can provide a readout of the electrostatic active site environment, an environment that is otherwise difficult to assess.