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Thioredoxin/glutathione reductase from Schistosoma mansoni (SmTGR) is a multifunctional enzyme that catalyzes the reduction of glutathione (GSSG) and thioredoxin, as well as the deglutathionylation of peptide and non-peptide substrates. SmTGR structurally resembles known glutathione reductases (GR) and thioredoxin reductases (TrxR) but with an appended N-terminal domain that has a typical glutaredoxin (Grx) fold. Despite structural homology with known GRs, the site of GSSG reduction has frequently been reported as the Grx domain, based primarily on aerobic, steady-state kinetic measurements and x-ray crystallography. Here, we present an anaerobic characterization of a series of variant SmTGRs to establish the site of GSSG reduction as the cysteine pair most proximal to the FAD, Cys154/Cys159, equivalent to the site of GSSG reduction in GRs. Anaerobic steady-state analysis of U597C, U597S, U597C + C31S, and I592STOP SmTGR demonstrate that the Grx domain is not involved in the catalytic reduction of GSSG, as redox silencing of the C-terminus results in no modulation of the observed turnover number (∼0.025 s−1) and redox silencing of the Grx domain results in an increased observed turnover number (∼0.08 s−1). Transient-state single turnover analysis of these variants corroborates this, as the slowest rate observed titrates hyperbolically with GSSG concentration and approaches a limit that coincides with the respective steady-state turnover number for each variant. Numerical integration fitting of the transient state data can only account for the observed trends when competitive binding of the C-terminus is included, indicating that the partitioning of electrons to either substrate occurs at the Cys154/Cys159 disulfide rather than the previously proposed Cys596/Sec597 sulfide/selenide. Paradoxically, truncating the C-terminus at Ile592 results in a loss of GR activity, indicating a crucial non-redox role for the C-terminus. 

Flavin disulfide reductases (FDRs) are FAD-dependent enzymes that transmit electrons from NAD(P)H to reduce specific oxidant substrate disulfides. These enzymes have been studied extensively, most particularly the paradigm examples: glutathione reductase and thioredoxin reductase. The common, though not universal, traits of the family include a tyrosine- or phenylalanine-gated binding pocket for NAD(P) nicotinamides adjacent to the FAD isoalloxazine re-face, and a disulfide stacked against the si-face of the isoalloxazine whose dithiol form is activated for subsequent exchange reactions by a nearby histidine acting as a base. This arrangement promotes transduction of the reducing equivalents for disulfide exchange relay reactions. From an observational standpoint the proximal parallel stacking of three redox moieties induces up to three opportunities for unique charge transfer interactions (NAD(P)H FAD, NAD(P)+•FADH2, and FAD•thiolate). In transient state, the charge transfer transitions provide discrete signals to assign reaction sequences. This review summarizes the lineage of observations for the FDR enzymes that have been extensively studied. Where applicable and in order to chart a consistent interpretation of the record, only data derived from studies that used anaerobic methods are cited. These data reveal a recurring theme for catalysis that is elaborated with specific additional functionalities for each oxidant substrate. 

Seafloor moorings measuring pressure and temperature were deployed from April to September 2023 at three sites near the route of the fiber optic telecommunications cable that extends offshore of Oliktok Point, Alaska. The raw data data (1 Hertz (Hz) sampling) are processed for hourly estimates of the ocean surface wave conditions, along with average seawater temperature and average depth. The sites were ice-covered from April to July, then mostly open water in August and September. The data were collected to calibrate proxy wave measurements using Distributed Acoustic Sensing (DAS) from the telecommunications cable. 

Abstract. The melt of snow and sea ice during the Arctic summer is a significant source of relatively fresh meltwater in the central Arctic. The fate of this freshwater – whether in surface melt ponds, or thin layers underneath the ice and in leads – impacts atmosphere-ice-ocean interactions and their subsequent coupled evolution. Here, we combine analyses of datasets from the Multidisciplinary drifting Observatory for the Study of Arctic Climate (MOSAiC) expedition (June–July, 2020) to understand the key drivers of the sea ice freshwater budget in the Central Arctic and the fate of this water over time. Freshwater budget analyses suggest that a relatively high fraction (58 %) is derived from surface melt. Additionally, the contribution from stored precipitation (snowmelt) significantly outweighs by five times the input from in situ summer precipitation (rain). The magnitude and rate of local meltwater production are remarkably similar to that observed on the prior Surface Heat Budget of the Arctic Ocean (SHEBA) campaign. A relatively small fraction (10 %) of freshwater from melt remains in ponds, which is higher on more deformed second-year ice compared to first-year ice later in the summer. Most meltwater drains via lateral and vertical drainage channels, with vertical drainage enabling storage of freshwater internally in the ice by freshening of brine channels. In the upper ocean, freshwater can accumulate in transient meltwater layers on the order of 10 cm to 1 m thick in leads and under the ice. The presence of such layers substantially impacts the coupled system by reducing bottom melt and allowing false bottom growth, reducing heat, nutrient and gas exchange, and influencing ecosystem productivity. Regardless, the majority fraction of freshwater from melt is inferred to be ultimately incorporated into upper ocean (75 %) or stored internally in the ice (14 %). Comparison of key source and sink terms with estimates from the CESM2 climate model suggest that simulated freshwater storage in melt ponds is dramatically underestimated. This suggests pond drainage terms should be investigated as a likely explanation. 

Thioredoxin/glutathione reductase (TGR) from the platyhelminthic parasitic worms has recently been identified as a drug target for the treatment of schistosomiasis. Schistosomes lack catalase, and so are heavily reliant on the regeneration of reduced thioredoxin (Trx) and glutathione (GSH) to reduce peroxiredoxins that ameliorate oxidative damage from hydrogen peroxide generated by the host immune response. This study focuses on the characterization of the catalytic mechanism ofSchistosoma mansoniTGR (SmTGR). Variant forms of SmTGR were studied to assign the function of residues that participate in the electron distribution chain within the enzyme. Using anaerobic transient state spectrophotometric methods, redox changes for the FAD and NADPH were observed and the function of specific residues was defined from observation of charge transfer absorption transitions that are indicative of specific complexations and redox states. The C159S variant prevented distribution of electrons beyond the flavin and as such did not accumulate thiolate-FAD charge transfer absorption. The lack of this absorption facilitated observation of a new charge transfer absorption consistent with proximity of NADPH and FAD. The C159S variant was used to confine electrons from NADPH at the flavin, and it was shown that NADPH and FAD exchange hydride in both directions and come to an equilibrium that yields only fractional FAD reduction, suggesting that both have similar reduction potentials. Mutation of U597 to serine resulted in sustained thiolate-FAD charge transfer absorption and loss of the ability to reduce Trx, indicating that the C596-U597 disulfide functions in the catalytic sequence to receive electrons from the C154 C159 pair and distribute them to Trx. No kinetic evidence for a loss or change in function associated with the distal C28-C31 disulfide was observed when the C31S variant reductive half-reaction was observed. The Y296A variant was shown to slow the rate of but increase extent of reduction of the flavin, and the dissociation of NADP⁺. The H571 residue was confirmed to be the residue responsible for the deprotonation of the C159 thiol, increasing its reactivity and generating the prominent thiolate-FAD charge transfer absorption that accumulates with oxidation of the flavin. 

Precise measurements of Arctic sea ice mass balance are necessary to understand the rapidly changing sea ice cover and its representation in climate models. During the Multidisciplinary drifting Observatory for the Study of Arctic Climate (MOSAiC) expedition, we made repeat point measurements of snow and ice thickness on primarily level first- and second-year ice (FYI, SYI) using ablation stakes and ice thickness gauges. This technique enabled us to distinguish surface and bottom (basal) melt and characterize the importance of oceanic versus atmospheric forcing. We also evaluated the time series of ice growth and melt in the context of other MOSAiC observations and historical mass balance observations from the Surface Heat Budget of the Arctic (SHEBA) campaign and the North Pole Environmental Observatory (NPEO). Despite similar freezing degree days, average ice growth at MOSAiC was greater on FYI (1.67 m) and SYI (1.23 m) than at SHEBA (1.45 m, 0.53 m), due in part to initially thinner ice and snow conditions on MOSAiC. Our estimates of effective snow thermal conductivity, which agree with SHEBA results and other MOSAiC observations, are unlikely to explain the difference. On MOSAiC, FYI grew more and faster than SYI, demonstrating a feedback loop that acts to increase ice production after multi-year ice loss. Surface melt on MOSAiC (mean of 0.50 m) was greater than at NPEO (0.18 m), with considerable spatial variability that correlated with surface albedo variability. Basal melt was relatively small (mean of 0.12 m), and higher than NPEO observations (0.07 m). Finally, we present observations showing that false bottoms reduced basal melt rates in some FYI cases, in agreement with other observations at MOSAiC. These detailed mass balance observations will allow further investigation into connections between the carefully observed surface energy budget, ocean heat fluxes, sea ice, and ecosystem at MOSAiC and during other campaigns. 

Thioredoxin/glutathione reductase from Schistosoma mansoni (SmTGR) catalyzes the reduction of both oxidized thioredoxin and glutathione with electrons from reduced nicotinamide adenine dinucleotide phosphate (NADPH). SmTGR is a drug target for the treatment of Schistosomiasis, an infection caused by Schistosoma platyhelminths residing in the blood vessels of the host. Schistosoma spp. are reliant on TGR enzymes as they lack catalase and so use reduced thioredoxin and glutathione to regenerate peroxiredoxins consumed in the detoxification of reactive oxygen species. SmTGR is a flavin adenine dinucleotide (FAD)-dependent enzyme, and we have used the flavin as a spectrophotometric reporter to observe the movement of electrons within the enzyme. The data show that NADPH fractionally reduces the active site flavin with an observed rate constant estimated in this study to be ∼3000 s-1. The flavin then reoxidizes by passing electrons at a similar rate to the proximal Cys159-Cys154 disulfide pair. The dissociation of NADP+ occurs with a rate of ∼180 s-1, which induces the deprotonation of Cys159, and this coincides with the accumulation of an intense FAD-thiolate charge transfer band. It is proposed that the electrons then pass to the Cys596-Cys597 disulfide pair of the associated subunit in the dimer with a net rate constant of ∼2 s-1. (Note: Cys597 is Sec597 in wild-type (WT) SmTGR.) From this position, the electrons can be passed to oxidized thioredoxin or further into the protein to reduce the Cys28-Cys31 disulfide pair of the originating subunit of the dimer. From the Cys28-Cys31 center, electrons can then pass to oxidized glutathione that has a binding site directly adjacent. 

Abstract We present results from wide-field imaging of the resolved stellar populations of the dwarf spheroidal galaxies Cassiopeia III (And XXXII) and Perseus I (And XXXIII), two satellites in the outer stellar halo of the Andromeda galaxy (M31). Our WIYN pODI photometry traces the red giant star population in each galaxy to ∼2.5−3 half-light radii from the galaxy center. We use the tip of the red giant branch (TRGB) method to derive distances of (m−M)₀= 24.62 ± 0.12 mag (839 ${}_{-45}^{+48}$kpc, or ${156}_{-13}^{+16}$kpc from M31) for Cas III and 24.47 ± 0.13 mag (738 ${}_{-45}^{+48}$kpc, or 351 ${}_{-16}^{+17}$kpc from M31) for Per I. These values are consistent within the errors with TRGB distances derived from a deeper Hubble Space Telescope study of the galaxies’ inner regions. For each galaxy, we derive structural parameters, total magnitude, and central surface brightness. We also place upper limits on the ratio of neutral hydrogen gas mass to optical luminosity, confirming the gas-poor nature of both galaxies. We combine our data set with corresponding data for the M31 satellite galaxy Lacerta I (And XXXI) from earlier work and search for substructure within the RGB star populations of Cas III, Per I, and Lac I. We find an overdense region on the west side of Lac I at a significance level of 2.5σ–3σand a low-significance filament extending in the direction of M31. In Cas III, we identify two modestly significant overdensities near the center of the galaxy and another at two half-light radii. Per I shows no evidence for substructure in its RGB star population, which may reflect this galaxy’s isolated nature. 

Dihydropyrimidine dehydrogenase (DPD) is a flavin dependent enzyme that catalyzes the reduction of the 5,6-vinylic bond of pyrimidines uracil and thymine with electrons from NADPH. DPD has two active sites that are separated by ∼60 Å. At one site NADPH binds adjacent to an FAD cofactor and at the other pyrimidine binds proximal to an FMN. Four Fe4S4 centers span the distance between these active sites. It has recently been established that the enzyme undergoes reductive activation prior to reducing the pyrimidine. In this initial process NADPH is oxidized at the FAD site and electrons are transmitted to the FMN via the Fe4S4 centers to yield the active state with a cofactor set of FAD•4(Fe4S4)•FMNH2. The catalytic chemistry of DPD can be studied in transient-state by observation of either NADPH consumption or charge transfer absorption associated with complexation of NADPH adjacent to the FAD. Here we have utilized both sets of absorption transitions to find evidence for specific additional aspects of the DPD mechanism. Competition for binding with NADP+ indicates that the two charge transfer species observed in activation/single turnover reactions arise from NADPH populating the FAD site before and after reductive activation. An additional charge transfer species is observed to accumulate at longer times when high NADPH concentrations are mixed with the enzyme•pyrimidine complex and this data can be modelled based on asymmetry in the homodimer. It was also shown that, like pyrimidines, dihydropyrimidines induce rapid reductive activation indicating that the reduced pyrimidine formed in turnover can stimulate the reinstatement of the active state of the enzyme. Investigation of the reverse reaction revealed that dihydropyrimidines alone can reductively activate the enzyme, albeit inefficiently. In the presence of dihydropyrimidine and NADP+ DPD will form NADPH but apparently without measurable reductive activation. Pyrimidines that have 5-substituent halogens were utilized to probe both reductive activation and turnover. The linearity of the Hammett plot based on the rate of hydride transfer to the pyrimidine establishes that, at least to the radius of an iodo-group, the 5-substituent volume does not have influence on the observed kinetics of pyrimidine reduction. 

The microstructure of the uppermost portions of a melting Arctic sea ice cover has a disproportionately large influence on how incident sunlight is reflected and absorbed in the ice/ocean system. The surface scattering layer (SSL) effectively backscatters solar radiation and keeps the surface albedo of melting ice relatively high compared to ice with the SSL manually removed. Measurements of albedo provide information on how incoming shortwave radiation is partitioned by the SSL and have been pivotal to improving climate model parameterizations. However, the relationship between the physical and optical properties of the SSL is still poorly constrained. Until now, radiative transfer models have been the only way to infer the microstructure of the SSL. During the MOSAiC expedition of 2019–2020, we took samples and, for the first time, directly measured the microstructure of the SSL on bare sea ice using X-ray micro-computed tomography. We show that the SSL has a highly anisotropic, coarse, and porous structure, with a small optical diameter and density at the surface, increasing with depth. As the melting surface ablates, the SSL regenerates, maintaining some aspects of its microstructure throughout the melt season. We used the microstructure measurements with a radiative transfer model to improve our understanding of the relationship between physical properties and optical properties at 850 nm wavelength. When the microstructure is used as model input, we see a 10–15% overestimation of the reflectance at 850 nm. This comparison suggests that either a) spatial variability at the meter scale is important for the two in situ optical measurements and therefore a larger sample size is needed to represent the microstructure or b) future work should investigate either i) using a ray-tracing approach instead of explicitly solving the radiative transfer equation or ii) using a more appropriate radiative transfer model.