Heparin treatment mitigates radiation-induced oral mucositis in mice by interplaying with repopulation processes.

PURPOSE: To investigate the mechanistic background of the muco-protective effect of systemic heparin treatment on the development of radiation-induced oral mucositis in mice. MATERIALS AND METHODS: Fractionated irradiation was given to the snouts of male C3H/Neu mice over 2 weeks (10â€¯× 3 Gy), either alone or in combination with daily subcutaneous application of unfractionated or low molecular weight heparin (40 or 200 I.U./mouse, respectively). Over this course of 14 days, groups of mice (n = 3) were sacrificed every second day, their tongues excised and processed for histological analysis. The epithelial radiation response with and without heparin treatment was evaluated in terms of tissue morphology, proliferation and expression of cell contact molecules. RESULTS: Systemic treatment with heparins significantly reduced the cellular effects of irradiation to the oral epithelium. Heparin treated animals showed significantly higher total epithelial cell numbers and thickness throughout the study course. Bromodeoxyuridine (BrdU) incorporation analyses revealed that markedly more epithelial cells retained their proliferative capacity in the beginning of the first treatment week, but the proliferation of the mucosa was not stimulated during the rest of the study course. The expression of the adherens junction protein ß­catenin was slightly elevated in heparin treated animals, on day 2 the increase was statistically significant. The expression of e­cadherin and occludin was mostly unaffected by the concomitant heparin treatment. CONCLUSION: The findings of this study indicate an interplay of additional heparin treatment with the repopulation processes, leading to an earlier onset of this adaptive radiation response in oral mucosa. Importantly, we could demonstrate that the protective potential of heparin did not rely on stimulation of normal tissue proliferation. Since both heparin preparations are already approved for clinical use, they are considered as promising candidates for future clinical studies.

Recommendations for the forensic diagnosis of sex and age from skeletons.

In both diagnostic fields a two-stage strategy is recommended: to first use "field" methods that are quick and easy but more imprecise and then "laboratory" methods that are time consuming but more precise. In preparing skeletal work, individuality of a skeleton should be checked, traces of diseases sought and time since death assessed. For sexing non-adults, the field methods are tooth mineralisation, long bone length and a few morphological skull and pelvis characteristics, for adults it is the morphology of pelvis and skull, and for both age groups the advanced laboratory method is molecular biology. For ageing non-adults the methods are mineralisation of teeth, long bone length and epiphysis development. For ageing adults the advanced laboratory method is aspartic acid racemisation. Less accurate laboratory methods are cement ring counts and histology of bones and teeth. Quick morphological methods using the pubic symphysis and other traits in combinations follow. Finally, cranial sutures and tooth number give a quick and rough impression. For the selection of a method and the assessment of its value the stochastic error produced for the reference sample is the decisive criterion; it should also be used to assess the reliability of a single diagnosis. Prerequisites for all work with skeletons are not only a complete knowledge of the relevant biology as well as specific techniques but also initial detailed instructions and with forensic applications, personal experience.

A subsequent fit of time series and amplitude histogram of patch-clamp records reveals rate constants up to 1 per microsecond.

Fast gating in time series of patch-clamp current demands powerful tools to reveal the rate constants of the adequate Hidden Markov model. Here, two approaches are presented to improve the temporal resolution of the direct fit of the time series. First, the prediction algorithm is extended to include intermediate currents between the nominal levels as caused by the anti-aliasing filter. This approach can reveal rate constants that are about 4 times higher than the corner frequency of the anti-aliasing filter. However, this approach is restricted to time series with very low noise. Second, the direct fit of the time series is combined with a beta fit, i.e., a fit of the deviations of the amplitude histogram from the Gaussian distribution. Since the "theoretical" amplitude histograms for higher-order Bessel filters cannot be calculated by analytical tools, they are generated from simulated time series. In a first approach, a simultaneous fit of the time series and of the Beta fit is tested. This simultaneous fit, however, inherits the drawbacks of both approaches, not the benefits. More successful is a subsequent fit: The fit of the time series yields a set of rate constants. The subsequent Beta fit uses the slow rate constants of the fit of the time series as fixed parameters and the optimization algorithm is restricted to the fast ones. The efficiency of this approach is illustrated by means of time series obtained from simulation and from the dominant K+ channel in Chara. This shows that temporal resolution can reach the microsecond range.

Structure of alanine dehydrogenase from Archaeoglobus: active site analysis and relation to bacterial cyclodeaminases and mammalian mu crystallin.

The hyperthermophilic archaeon Archaeoglobus fulgidus contains an L-Ala dehydrogenase (AlaDH, EC 1.4.1.1) that is not homologous to known bacterial dehydrogenases and appears to represent a previously unrecognized archaeal group of NAD-dependent dehydrogenases. The gene (Genbank; TIGR AF1665) was annotated initially as an ornithine cyclodeaminase (OCD) on the basis of strong homology with the mu crystallin/OCD protein family. We report the structure of the NAD-bound AF1665 AlaDH (AF-AlaDH) at 2.3 A in a C2 crystal form with the 70 kDa dimer in the asymmetric unit, as the first structural representative of this family. Consistent with its lack of homology to bacterial AlaDH proteins, which are mostly hexameric, the archaeal dimer has a novel structure. Although both types of AlaDH enzyme include a Rossmann-type NAD-binding domain, the arrangement of strands in the C-terminal half of this domain is novel, and the other (catalytic) domain in the archaeal protein has a new fold. The active site presents a cluster of conserved Arg and Lys side-chains over the pro-R face of the cofactor. In addition, the best ordered of the 338 water molecules in the structure is positioned well for mechanistic interaction. The overall structure and active site are compared with other dehydrogenases, including the AlaDH from Phormidium lapideum. Implications for the catalytic mechanism and for the structures of homologs are considered. The archaeal AlaDH represents an ancient and previously undescribed subclass of Rossmann-fold proteins that includes bacterial ornithine and lysine cyclodeaminases, marsupial lens proteins and, in man, a thyroid hormone-binding protein that exhibits 30% sequence identity with AF1665.

Distributions-per-level: a means of testing level detectors and models of patch-clamp data.

Level or jump detectors generate the reconstructed time series from a noisy record of patch-clamp current. The reconstructed time series is used to create dwell-time histograms for the kinetic analysis of the Markov model of the investigated ion channel. It is shown here that some additional lines in the software of such a detector can provide a powerful new means of patch-clamp analysis. For each current level that can be recognized by the detector, an array is declared. The new software assigns every data point of the original time series to the array that belongs to the actual state of the detector. From the data sets in these arrays distributions-per-level are generated. Simulated and experimental time series analyzed by Hinkley detectors are used to demonstrate the benefits of these distributions-per-level. First, they can serve as a test of the reliability of jump and level detectors. Second, they can reveal beta distributions as resulting from fast gating that would usually be hidden in the overall amplitude histogram. Probably the most valuable feature is that the malfunctions of the Hinkley detectors turn out to depend on the Markov model of the ion channel. Thus, the errors revealed by the distributions-per-level can be used to distinguish between different putative Markov models of the measured time series.

Comparison between the nitric oxide reductase family and its aerobic relatives, the cytochrome oxidases.

The denitrification pathway has been studied in the hyperthermophilic archaeon Pyrobaculum aerophilum. In contrast with Gram-negative bacteria, all four denitrification enzymes are membrane-bound. P. aerophilum is also the only denitrifyer identified so far in which menaquinol is the electron donor to all four denitrification reductases. The NO reductase (NOR) of P. aerophilum belongs to the superfamily of haem-copper oxidases and is of the qNOR (quinol-dependent) type. Three types of NOR have been purified so far: cNOR (cytochrome c/pseudoazurin-dependent), qNOR and qCu(A)NOR (qNOR that contains Cu(A) at the electron entry site). It is proposed that the NORs and the various cytochrome oxidases have evolved by modular evolution, in view of the structure of their electron donor sites. qNOR is further proposed to be the ancestor of all NORs and cytochrome oxidases belonging to the superfamily of haem-copper oxidases.

Properties of a thermostable nitrate reductase from the hyperthermophilic archaeon Pyrobaculum aerophilum.

The nitrate reductase of the hyperthermophilic archaeon Pyrobaculum aerophilum was purified 137-fold from the cytoplasmic membrane. Based on sodium dodecyl sulfate-polyacrylamide gel electrophoresis analysis, the enzyme complex consists of three subunits with apparent molecular weights of 130,000, 52,000, and 32,000. The enzyme contained molybdenum (0.8-mol/mol complex), iron (15.4-mol/mol complex) and cytochrome b (0.49-mol/mol complex) as cofactors. The P. aerophilum nitrate reductase distinguishes itself from nitrate reductases of mesophilic bacteria and archaea by its very high specific activity using reduced benzyl viologen as the electron donor (V(max) with nitrate, 1,162 s(-1) (326 U/mg); V(max) with chlorate, 1,348 s(-1) (378 U/mg) [assayed at 75 degrees C]). The K(m) values for nitrate and chlorate were 58 and 140 microM, respectively. Azide was a competitive inhibitor and cyanide was a noncompetitive inhibitor of the nitrate reductase activity. The temperature optimum for activity was > 95 degrees C. When incubated at 100 degrees C, the purified nitrate reductase had a half-life of 1.5 h. This study constitutes the first description of a nitrate reductase from a hyperthermophilic archaeon.

Crystal structures of a novel ferric reductase from the hyperthermophilic archaeon Archaeoglobus fulgidus and its complex with NADP+.

BACKGROUND: Studies performed within the last decade have indicated that microbial reduction of Fe(III) to Fe(II) is a biologically significant process. The ferric reductase (FeR) from Archaeoglobus fulgidus is the first reported archaeal ferric reductase and it catalyzes the flavin-mediated reduction of ferric iron complexes using NAD(P)H as the electron donor. Based on its catalytic activity, the A. fulgidus FeR resembles the bacterial and eukaryotic assimilatory type of ferric reductases. However, the high cellular abundance of the A. fulgidus FeR (approximately 0.75% of the total soluble protein) suggests a catabolic role for this enzyme as the terminal electron acceptor in a ferric iron-based respiratory pathway [1]. RESULTS: The crystal structure of recombinant A. fulgidus FeR containing a bound FMN has been solved at 1.5 A resolution by multiple isomorphous replacement/ anomalous diffraction (MIRAS) phasing methods, and the NADP+- bound complex of FeR was subsequently determined at 1.65 A resolution. FeR consists of a dimer of two identical subunits, although only one subunit has been observed to bind the redox cofactors. Each subunit is organized around a six-stranded antiparallel beta barrel that is homologous to the FMN binding protein from Desulfovibrio vulgaris. This fold has been shown to be related to a circularly permuted version of the flavin binding domain of the ferredoxin reductase superfamily. The A. fulgidus ferric reductase is further distinguished from the ferredoxin reductase superfamily by the absence of a Rossmann fold domain that is used to bind the NAD(P)H. Instead, FeR uses its single domain to provide both the flavin and the NAD(P)H binding sites. Potential binding sites for ferric iron complexes are identified near the cofactor binding sites. CONCLUSIONS: The work described here details the structures of the enzyme-FMN, enzyme-FMN-NADP+, and possibly the enzyme-FMN-iron intermediates that are present during the reaction mechanism. This structural information helps identify roles for specific residues during the reduction of ferric iron complexes by the A. fulgidus FeR.

A novel copper A containing menaquinol NO reductase from Bacillus azotoformans.

The molecular biology and biochemistry of denitrification in gram-negative bacteria has been studied extensively. However, little is known about this process in gram-positive bacteria. We have purified the NO reductase from the cytoplasmic membrane of the gram-positive bacterium Bacillus azotoformans. The purified enzyme consists of two subunits with apparent molecular masses of 16 and 40 kDa based on SDS-PAGE. Analytical and spectroscopic determinations revealed the presence of one non-heme iron, two copper atoms and of two b-type hemes per enzyme complex. Heme c was absent. Using EPR and UV-visible spectroscopy, it was determined that one of the hemes is a low-spin heme b, in which the two axial histidine imidazole planes are positioned at an angle of 60-70 degrees. The second heme b is high-spin binding CO in the reduced state. The high-spin heme center and the non-heme iron are EPR silent. They are proposed to form a binuclear center where reduction of NO occurs. There are two novel features of this enzyme that distinguish it from other NO reductases. First, the enzyme contains copper in form of copper A, an electron carrier up to now only detected in cytochrome oxidases and nitrous oxide reductases. Second, the enzyme uses menaquinol as electron donor, whereas cytochrome c, which is the substrate of other NO reductases, is not used. Copper A and both hemes are reducible by menaquinol. This new NO reductase is thus a menaquinol:NO oxidoreductase. With respect to its prosthetic groups the B. azotoformans NO reductase is a true hybrid between copper A containing cytochrome oxidases and NO reductases present in gram-negative bacteria. It may represent the most ancient "omnipotent" progenitor of the family of heme-copper oxidases.

Analyzing your complexes: structure of the quinol-fumarate reductase respiratory complex.

The integral membrane protein complex quinol-fumarate reductase catalyzes the terminal step of a major anaerobic respiratory pathway. The homologous enzyme succinate-quinone oxidoreductase participates in aerobic respiration both as complex II and as a member of the Krebs cycle. Last year, two structures of quinol-fumarate reductases were reported. These structures revealed the cofactor organization linking the fumarate and quinol sites, and showed a cofactor arrangement across the membrane that is suggestive of a possible energy coupling function.

Lipopolysaccharides of Rhizobium etli strain G12 act in potato roots as an inducing agent of systemic resistance to infection by the cyst nematode Globodera pallida.

Recent studies have shown that living and heat-killed cells of the rhizobacterium Rhizobium etli strain G12 induce in potato roots systemic resistance to infection by the potato cyst nematode Globodera pallida. To better understand the mechanisms of induced resistance, we focused on identifying the inducing agent. Since heat-stable bacterial surface carbohydrates such as exopolysaccharides (EPS) and lipopolysaccharides (LPS) are essential for recognition in the symbiotic interaction between Rhizobium and legumes, their role in the R. etli-potato interaction was studied. EPS and LPS were extracted from bacterial cultures, applied to potato roots, and tested for activity as an inducer of plant resistance to the plant-parasitic nematode. Whereas EPS did not affect G. pallida infection, LPS reduced nematode infection significantly in concentrations as low as 1 and 0.1 mg ml(-1). Split-root experiments, guaranteeing a spatial separation of inducing agent and challenging pathogen, showed that soil treatments of one half of the root system with LPS resulted in a highly significant (up to 37%) systemic induced reduction of G. pallida infection of potato roots in the other half. The results clearly showed that LPS of R. etli G12 act as the inducing agent of systemic resistance in potato roots.

Identification and characterization of a novel ferric reductase from the hyperthermophilic Archaeon Archaeoglobus fulgidus.

Archaeoglobus fulgidus, a hyperthermophilic sulfate-reducing Archaeon, contains high Fe(3+)-EDTA reductase activity in its soluble protein fraction. The corresponding enzyme, which constitutes about 0.75% of the soluble protein, was purified 175-fold to homogeneity. Based on SDS-polyacrylamide gel electrophoresis, the ferric reductase consists of a single subunit with a M(r) of 18,000. The M(r) of the native enzyme was determined by size exclusion chromatography to be 40,000 suggesting that the native ferric reductase is a homodimer. The enzyme uses both NADH and NADPH as electron donors to reduce Fe(3+)-EDTA. Other Fe(3+) complexes and dichlorophenolindophenol serve as alternative electron acceptors, but uncomplexed Fe(3+) is not utilized. The purified enzyme strictly requires FMN or FAD as a catalytic intermediate for Fe(3+) reduction. Ferric reductase also reduces FMN and FAD, but not riboflavin, with NAD(P)H which classifies the enzyme as a NAD(P)H:flavin oxidoreductase. The enzyme exhibits a temperature optimum of 88 degrees C. When incubated at 85 degrees C, the enzyme activity half-life was 2 h. N-terminal sequence analysis of the purified ferric reductase resulted in the identification of the hypothetical gene, AF0830, of the A. fulgidus genomic sequence. The A. fulgidus ferric reductase shares amino acid sequence similarity with a family of NAD(P)H:FMN oxidoreductases but not with any ferric reductases suggesting that the A. fulgidus ferric reductase is a novel enzyme.

An Escherichia coli mutant quinol:fumarate reductase contains an EPR-detectable semiquinone stabilized at the proximal quinone-binding site.

The EPR and thermodynamic properties of semiquinone (SQ) species stabilized by mammalian succinate:quinone reductase (SQR) in situ in the mitochondrial membrane and in the isolated enzyme have been well documented. The equivalent semiquinones in bacterial membranes have not yet been characterized, either in SQR or quinol:fumarate reductase (QFR) in situ. In this work, we describe an EPR-detectable QFR semiquinone using Escherichia coli mutant QFR (FrdC E29L) and the wild-type enzyme. The SQ exhibits a g = 2.005 signal with a peak-to-peak line width of approximately 1.1 milliteslas at 150 K, has a midpoint potential (E(m(pH 7.2))) of -56.6 mV, and has a stability constant of approximately 1.2 x 10(-2) at pH 7.2. It shows extremely fast spin relaxation behavior with a P(1/2) value of >>500 milliwatts at 150 K, which closely resembles the previously described SQ species (SQ(s)) in mitochondrial SQR. This SQ species seems to be present also in wild-type QFR, but its stability constant is much lower, and its signal intensity is near the EPR detection limit around neutral pH. In contrast to mammalian SQR, the membrane anchor of E. coli QFR lacks heme; thus, this prosthetic group can be excluded as a spin relaxation enhancer. The trinuclear iron-sulfur cluster FR3 in the [3Fe-4S](1+) state is suggested as the dominant spin relaxation enhancer of the SQ(FR) spins in this enzyme. E. coli QFR activity and the fast relaxing SQ species observed in the mutant enzyme are sensitive to the inhibitor 2-n-heptyl-4-hydroxyquinoline N-oxide (HQNO). In wild-type E. coli QFR, HQNO causes EPR spectral line shape perturbations of the iron-sulfur cluster FR3. Similar spectral line shape changes of FR3 are caused by the FrdC E29L mutation, without addition of HQNO. This indicates that the SQ and the inhibitor-binding sites are located in close proximity to the trinuclear iron-sulfur cluster FR3. The data further suggest that this site corresponds to the proximal quinone-binding site in E. coli QFR.

Methylcobalamin:homocysteine methyltransferase from Methanobacterium thermoautotrophicum. Identification as the metE gene product.

Methanobacterium thermoautotrophicum is a methane-forming archaeon that grows on H2 and CO2 as sole carbon and energy source. Cell extracts of the methanogen were found to contain methylcobalamin: homocysteine methyltransferase activity which was purified 3000-fold to a specific activity of approximately 500 U.mg-1 protein. SDS/PAGE revealed the presence of a polypeptide with an apparent molecular mass of 34 kDa. Via its N-terminal amino acid sequence, the 34-kDa polypeptide was identified as the metE gene product. The metE gene was heterologously expressed in Escherichia coli. The overproduced protein was recovered in the inclusion body fraction and was found to be inactive. The protein could be partially solubilized by unfolding in 8 M urea and then refolding. The solubilized protein had a specific activity of 450 U.mg-1. It exhibited first-order kinetics with respect to methylcobalamin concentration and Michaelis-Menten kinetics with respect to L-homocysteine concentration (apparent Km 0.1 mM). The enzyme was specific for L-homocysteine as methyl acceptor. Methylcobalamin could be substituted with methylcobinamide as methyl donor.

Effect of tungstate on nitrate reduction by the hyperthermophilic archaeon pyrobaculum aerophilum

Pyrobaculum aerophilum, a hyperthermophilic archaeon, can respire either with low amounts of oxygen or anaerobically with nitrate as the electron acceptor. Under anaerobic growth conditions, nitrate is reduced via the denitrification pathway to molecular nitrogen. This study demonstrates that P. aerophilum requires the metal oxyanion WO42- for its anaerobic growth on yeast extract, peptone, and nitrate as carbon and energy sources. The addition of 1 &mgr;M MoO42- did not replace WO42- for the growth of P. aerophilum. However, cell growth was completely inhibited by the addition of 100 &mgr;M MoO42- to the culture medium. At lower tungstate concentrations (0.3 &mgr;M and less), nitrite was accumulated in the culture medium. The accumulation of nitrite was abolished at higher WO42- concentrations (<0.7 &mgr;M). High-temperature enzyme assays for the nitrate, nitrite, and nitric oxide reductases were performed. The majority of all three denitrification pathway enzyme activities was localized to the cytoplasmic membrane, suggesting their involvement in the energy metabolism of the cell. While nitrite and nitric oxide specific activities were relatively constant at different tungstate concentrations, the activity of nitrate reductase was decreased fourfold at WO42- levels of 0.7 &mgr;M or higher. The high specific activity of the nitrate reductase enzyme observed at low WO42- levels (0.3 &mgr;M or less) coincided with the accumulation of nitrite in the culture medium. This study documents the first example of the effect of tungstate on the denitrification process of an extremely thermophilic archaeon. We demonstrate here that nitrate reductase synthesis in P. aerophilum occurs in the presence of high concentrations of tungstate.

NarL dimerization? Suggestive evidence from a new crystal form.

The structure of the Escherichia coli response regulator NarL has been solved in a new, monoclinic space group, and compared with the earlier orthorhombic crystal structure. Because the monoclinic crystal has two independent NarL molecules per asymmetric unit, we now have three completely independent snapshots of the NarL molecule: two from the monoclinic form and one from the orthorhombic. Comparison of these three structures shows the following: (a) The pairing of N and C domains of the NarL molecule proposed from the earlier analysis is in fact correct, although the polypeptide chain connecting domains was, and remains, disordered and not completely visible. The new structure exhibits identical relative orientation of N and C domains, and supplies some of the missing residues, leaving a gap of only seven amino acids. (b) Examination of corresponding features in the three independent NarL molecules shows that deformations in structure produced by crystal packing are negligible. (c) The "telephone receiver" model of NarL activation is confirmed. The N domain of NarL blocks the binding of DNA to the C domain that would be expected from the helix-turn-helix structure of the C domain. Hence, binding can only occur after significant displacement of N and C domains. (d) NarL monomers have a strong tendency toward dimerization involving contacts between helixes alpha 1 in the two monomers, and this may have mechanistic significance in DNA binding. Analogous involvement of helix alpha 1 in intermolecular contacts is also found in UhpA and in the CheY/CheZ complex.

Inhibition of human and duck hepatitis B virus by 2',3'-dideoxy-3'-fluoroguanosine in vitro.

The fluorinated guanosine analog 2',3'-dideoxy-3'-fluoroguanosine (FLG) has been shown to have an effect on duck hepatitis B virus (DHBV) in vivo and in vitro. In this study the inhibitory effect of FLG on DHBV and human hepatitis B virus (HBV) was evaluated in vitro. Cell lines transfected either with DHBV or HBV DNA and primary duck hepatocyte cell cultures were used. Virus production was analysed by PCR and a quantitative PCR was established for DHBV for determination of the inhibitory concentrations of the drug. 50% inhibition was achieved with an FLG concentration of 0.2 microg/ml (0.7 microM) and 90% inhibition was observed with an FLG concentration of 1.0 microg/ml (3.7 microM) using the DHBV transfected cell line. FLG showed an effect on DHBV production in primary duck hepatocyte cell cultures at concentrations down to 0.1 microg/ml (0.4 microM). However, the DHBV production returned to pre-treatment levels within a few days after cessation of treatment. HBV production in transfected cell lines was also inhibited by FLG. Both DHBV and HBV DNA-polymerases were inhibited by FLG triphosphate and 50% inhibition was observed at a concentration of 0.05 microg/ml (0.1 microM) for DHBV and 0.03 microg/ml (0.05 microM) for HBV. FLG is an efficient inhibitor of DHBV replication both in vivo and in vitro and of HBV in vitro which makes it a good candidate for treatment of HBV infections. However, it does not completely eliminate the virus since a relapse in virus production was observed when treatment was withdrawn. Therefore it would be interesting to evaluate FLG in combination with other types of anti-HBV drugs.

Purification and characterization of the selenate reductase from Thauera selenatis.

Thauera selenatis is one of two isolated bacterial species that can obtain energy by respiring anaerobically with selenate as the terminal electron acceptor. The reduction of selenate to selenite is catalyzed by a selenate reductase, previously shown to be located in the periplasmic space of the cell. This study describes the purification of the enzyme from T. selenatis grown anaerobically with selenate. The enzyme is a trimeric alphabetagamma complex with an apparent Mr of 180,000. The alpha, beta, and gamma subunits are 96 kDa, 40 kDa, and 23 kDa, respectively, in size. The selenate reductase contains molybdenum, iron, and acid-labile sulfur as prosthetic group constituents. UV-visible absorption spectroscopy also revealed the presence of one cytochrome b per alphabetagamma complex. The Km for selenate was determined to be 16 microM, and the Vmax was 40 micromol/min/mg of protein. The enzyme is specific for the reduction of selenate; nitrate, nitrite, chlorate, and sulfate were not reduced at detectable rates. These studies constitute the first description of a selenate reductase, which represents a new class of enzymes. The significance of this enzyme in relation to cell growth and energy generation is discussed.

Anthropometrical measurements and androgen levels in males, females, and hormonally untreated female-to-male transsexuals.

To elucidate the relationship between body build, androgens, and transsexual gender identity, anthropometric measurements were assessed in 15 hormonally untreated female-to-male-transsexuals (FMT). Nineteen healthy women (CF) (X = 22 years; 2 months), and 21 healthy men (CM) (X = 23; 7) were enrolled as controls. Baseline levels of testosterone (T; ng/dl), androstenedione (A4; ng/dl), dehydroepiandrosterone sulfate (DHEAS; ng/ml), and sex-hormone binding globulin (SHBG; microgram/ml) were assessed in 12 FMT, 15 CF, and in all CM. No control was under hormonal medication (including contraceptives). Absolute measurements in FMT were in accordance with their biological sex: they showed only small differences from the CF. However, FMT differed from CF in 7 of 14 sex-dimorphic indices of masculinity/femininity in body build. Of these 14 indices, 9 did not show a difference between FMT and CM. Hence, FMT presented a more masculine body build, particularly in fat distribution and bone proportions. Levels of T and A4 were significantly higher in FMT than in CF (T: 54.0 +/- 13.8 vs. 41.1 +/- 12.8; A4: 244.8 +/- 73.0 vs. 190.5 +/- 49.3), while DHEAS was higher in CM (3335 +/- 951) than in CF (2333 +/- 793) and in FMT (2679 +/- 1089). Altogether, 83.3% of FMT and 33.3% of CF were above normal values for at least one measured androgen. SHBG in FMT (1.21 +/- 0.70) and CF (1.87 +/- 0.91) was higher than in CM (0.49 +/- 0.18) and tended to be higher in CF than in FMT. Unbound T (T/SHBG ratio) was higher in FMT (72.0 +/- 67.6) than in CF (26.4 +/- 15.1) and correlated positively with manly body shape. Findings are discussed in relation to etiology of transsexualism.