Biomedical subjects

D P Baker

Publications and source records attributed to D P Baker.

At least 37 records · Page 2Linked to original sources

PubMed1997

Fibronectin type III repeats mediate RGD-independent adhesion and signaling through activated beta1 integrins.

Many cell-surface and extracellular matrix proteins contain multiple modular domains known as fibronectin type III (FNIII) repeats. Cells adhere to the extracellular matrix proteins fibronectin and tenascin in part by the interaction of certain integrins with the Arg-Gly-Asp (RGD) sequence, displayed on specific FNIII repeats. We have found that, after experimental activation of beta1 integrins, a number of cell types adhere and spread on FNIII repeats lacking RGD, derived from extracellular matrix proteins and cytokine receptors. Interaction between individual FNIII domains and beta1 integrins mediates focal adhesion kinase phosphorylation and subsequent stress fiber and focal contact formation. These data suggest that many FNIII-containing proteins may bind and signal through activated beta1 integrins, dramatically expanding the potential for integrin-dependent intercellular and cell-matrix communication.

Animals↗

PubMed1997

The COBE Spectra cell separator is more effective than the Haemonetics MCS-3P cell separator for peripheral blood progenitor cell harvest after mobilization with cyclophosphamide and filgrastim.

BACKGROUND: Peripheral blood is rapidly replacing bone marrow as a source of hematopoietic progenitor cells for autologous transplantation. The advantages of peripheral blood progenitor cell transplantation are enhanced by the ability to collect sufficient progenitor cells to ensure rapid neutrophil and platelet recovery in a single procedure on some cell separators. STUDY DESIGN AND METHODS: A prospective randomized study was undertaken to compare peripheral blood progenitor cell yields from two cell separators (MCS-3P, Haemonetics and Spectra, COBE). Fifteen consecutive patients were mobilized with cyclophosphamide 2 g per m2 (Day 0) and filgrastim 10 micrograms per kg (Days 1-11). Consecutive collections (Day 10, Day 11) were performed with each machine once: patients were randomly assigned to either machine for the initial collection. RESULTS: Collection time was longer on the MCS-3P (p = 0.001), and the volume processed was greater with the Spectra (p < 0.0001). Despite similar nucleated cell yield (p = 0.62), the yield of CD34+ cells (p = 0.001) and colony-forming units-granulocytic-monocytic (p = 0.0001) was significantly higher with the Spectra. The yield of nucleated cells per unit of blood volume processed was higher for the MCS-3P (p = 0.0007), while the CD34+ cell yield (p = 1) and colony-forming units-granulocytic-monocytic yield (p = 1) per unit of blood volume processed were similar for the two machines. The collection of CD34+ cells at levels > 2 x 10(6) per kg (p = 0.063), 5 x 10(6) per kg (p = 0.031), and colony-forming units-granulocytic-monocytic > 1 x 10(5) per kg (p = 0.25) after a single collection was superior for the Spectra. CONCLUSION: The yield of progenitor cells after collection on the Spectra was superior to that achieved with the MCS-3P, because of the larger volume of blood processed per procedure. This would permit more patients to undergo only one collection.

Blood Volume↗

PubMed1996

Engineered complementation in Escherichia coli aspartate transcarbamoylase. Heterotropic regulation by quaternary structure stabilization.

Escherichia coli aspartate transcarbamoylase regulates pyrimidine biosynthesis by altering its activity homotropically in response to one of its substrates and heterotropically in response to nucleotide effectors. The mechanism of this regulation involves two structurally and functionally different forms of the enzyme, one with low activity and low affinity for substrates (T state) and the other with high activity and high affinity for substrates (R state). Heterotropic regulation may be due to the direct transmission of a regulatory "signal" between the regulatory site and the active site some 60 A away and/or may involve altering the relative stability of the two forms of the enzyme. By combining a T state-stabilized mutant version of the enzyme, previously thought to have a defect in a heterotropic transmission pathway, with a known R state-stabilized mutant enzyme, we were able to restore some properties of the wild-type enzyme. These data imply that the relative stabilization of the T and R states of the enzyme is in part responsible for the homotropic and heterotropic properties of aspartate transcarbamoylase and that direct pathways for transmission of the heterotropic signals are unlikely.

Aspartate Carbamoyltransferase↗

PubMed1996

Catabolic ornithine carbamoyltransferase of Pseudomonas aeruginosa. Importance of the N-terminal region for dodecameric structure and homotropic carbamoylphosphate cooperativity.

Pseudomonas aeruginosa has an anabolic (ArgF) and a catabolic (ArcB) ornithine carbamoyltransferase (OTCase). Despite extensive sequence similarities, these enzymes function unidirectionally in vivo. In the dodecameric catabolic OTCase, homotropic cooperativity for carbamoylphosphate strongly depresses the anabolic reaction; the residue Glu1O5 and the C-terminus are known to be essential for this cooperativity. When Glu1O5 and nine C-terminal amino acids of the catabolic OTCase were introduced, by in vitro genetic manipulation, into the closely related, trimeric, anabolic (ArgF) OTCase of Escherichia coli, the enzyme displayed Michaelis-Menten kinetics and no cooperativity was observed. This indicates that additional amino acid residues are required to produce homotropic cooperativity and a dodecameric assembly. To localize these residues, we constructed several hybrid enzymes by fusing, in vivo or in vitro, the E. coli argF gene to the P. aeruginosa arcB gene. A hybrid enzyme consisting of 101 N-terminal ArgF amino acids fused to 233 C-terminal ArcB residues and the reciprocal ArcB-ArgF hybrid were both trimers with little or no cooperativity. Replacing the seven N-terminal residues of the ArcB enzyme by the corresponding six residues of E. coli ArgF enzyme produced a dodecameric enzyme which showed a reduced affinity for carbamoylphosphate and an increase in homotropic cooperativity. Thus, the N-terminal amino acids of catabolic OTCase are important for interaction with carbamoylphosphate, but do not alone determine dodecameric assembly. Hybrid enzymes consisting of either 26 or 42 N-terminal ArgF amino acids and the corresponding C-terminal ArcB residues were both trimeric, yet they retained some homotropic cooperativity. Within the N-terminal ArcB region, a replacement of motif 28-33 by the corresponding ArgF segment destabilized the dodecameric structure and the enzyme existed in trimeric and dodecameric states, indicating that this region is important for dodecameric assembly. These findings were interpreted in the light of the three-dimensional structure of catabolic OTCase, which allows predictions about trimer-trimer interactions. Dodecameric assembly appears to require at least three regions: the N- and C-termini (which are close to each other in a monomer), residues 28-33 and residues 147-154. Dodecameric structure correlates with high carbamoylphosphate cooperativity and thermal stability, but some trimeric hybrid enzymes retain cooperativity, and the dodecameric Glu1O5-->Ala mutant gives hyperbolic carbamoylphosphate saturation, indicating that dodecameric structure is neither necessary nor sufficient to ensure cooperativity.

Allosteric Regulation↗

PubMed1996

The allosteric activator ATP induces a substrate-dependent alteration of the quaternary structure of a mutant aspartate transcarbamoylase impaired in active site closure.

Aspartate transcarbamoylase from Escherichia coli shows homotropic cooperativity for aspartate as well as heterotropic regulation by nucleotides. Structurally, it consists of two trimeric catalytic subunits and three dimeric regulatory subunits, each chain being comprised of two domains. Glu-50 and Ser-171 are involved in stabilizing the closed conformation of the catalytic chain. Replacement of Glu-50 or Ser-171 by Ala in the holoenzyme has been shown previously to result in marked decreases in the maximal observed specific activity, homotropic cooperativity, and affinity for aspartate (Dembowski NJ, Newton CJ, Kantrowitz ER, 1990, Biochemistry 29:3716-3723; Newton CJ, Kantrowitz ER, 1990, Biochemistry 29:1444-1451). We have constructed a double mutant enzyme combining both mutations. The resulting Glu-50/ser-171-->Ala enzyme is 9-fold less active than the Ser-171-->Ala enzyme, 69-fold less active than the Glu-50-->Ala enzyme, and shows 1.3-fold and 1.6-fold increases in the [S]0.5Asp as compared to the Ser-171-->Ala and Glu-50-->Ala enzymes, respectively. However, the double mutant enzyme exhibits some enhancement of homotropic cooperativity with respect to aspartate, relative to the single mutant enzymes. At subsaturating concentrations of aspartate, the Glu-50/Ser-171 -->Ala enzyme is activated less by ATP than either the Glu-50-->Ala or Ser-171-->Ala enzyme, whereas CTP inhibition is intermediate between that of the two single mutants. As opposed to the wild-type enzyme, the Glu-50/Ser-171 -->Ala enzyme is activated by ATP and inhibited by CTP at saturating concentrations of aspartate. Structural analysis of the Ser-171-->Ala and Glu-50/Ser-171-->Ala enzymes by solution X-ray scattering indicates that both mutants exist in the same T quaternary structure as the wild-type enzyme in the absence of ligands, and in the same R quaternary structure in the presence of saturating N-(phosphonoacetyl)-L-aspartate. However, saturating concentrations of carbamoyl phosphate and succinate are unable to convert a significant fraction of either mutant enzyme population to the R quaternary structure, as has been observed previously for the Glu-50-->Ala enzyme. The curves for both the Ser-171-->Ala and Glu-50/Ser-171-->Ala enzymes obtained in the presence of substoichiometric amounts of PALA are linear combinations of the two extreme T and R states. The structural consequences of nucleotide binding to these two enzymes were also investigated. Most surprisingly, the direction and amplitude of the effect of ATP upon the double mutant enzyme were shown to vary depending upon the substrate analogue used.

Adenosine Triphosphate↗

PubMed1996

Analyzing team performance: in the eye of the beholder?

The effect of team-member experience was examined in relation to perceptions of importance of teamwork behaviors during an analysis of team performance. Importance ratings were collected from military aircrews (i.e., two-member teams) from three distinct types of aircraft. Results indicated that when assessing team behavior importance, less experienced team members weighted difficulty of performing team behaviors more heavily, whereas more experienced team members weighted time spent performing team behaviors more heavily. Implications for analyzing team performance, with respect to the process of conducting such analyses and the type of information collected, are discussed.

Aerospace Medicine↗

PubMed1995

Brain mechanisms in human classical conditioning: a PET blood flow study.

Five healthy male subjects participated in a classical conditioning experiment, and positron emission tomography (PET) was used to compare regional cerebral blood flow before and after conditioning. The subjects participated in three different experimental phases. In the first (habituation) phase they listened to 24 repetitions of a tone with random intervals. In the second (acquisition) phase, the tone was paired with a brief shock to the wrist. In the third (extinction) phase, the tone was presented alone again. 15OPET scans were taken during the habituation and extinction phases. Because the habituation and extinction phases were similar, any difference in blood flow to the tones presented during extinction probably reflected conditioning that occurred during the acquisition phase. Statistical parametric mapping (SPM) analysis of the PET data showed significantly increased activation in the right hemisphere in the orbito-frontal cortex, dorsolateral prefrontal cortex, inferior and superior frontal cortices, and inferior and middle temporal corticies. The only activated areas in the left hemisphere were area 19 and the superior frontal cortex. The results are interpreted as evidence for the involvement of cortical areas in human classical conditioning.

Acoustic Stimulation↗

PubMed1995

Weakening of the interface between adjacent catalytic chains promotes domain closure in Escherichia coli aspartate transcarbamoylase.

Aspartate transcarbamoylase from Escherichia coli is a dodecameric enzyme consisting of two trimeric catalytic subunits and three dimeric regulatory subunits. Asp-100, from one catalytic chain, is involved in stabilizing the C1-C2 interface by means of its interaction with Arg-65 from an adjacent catalytic chain. Replacement of Asp-100 by Ala has been shown previously to result in increases in the maximal specific activity, homotropic cooperativity, and the affinity for aspartate (Baker DP, Kantrowitz ER, 1993, Biochemistry 32:10150-10158). In order to determine whether these properties were due to promotion of domain closure induced by the weakening of the C1-C2 interface, we constructed a double mutant version of aspartate transcarbamoylase in which the Asp-100-->Ala mutation was introduced into the Glu-50-->Ala holoenzyme, a mutant in which domain closure is impaired. The Glu-50/Asp-100-->Ala enzyme is fourfold more active than the Glu-50-->Ala enzyme, and exhibits significant restoration of homotropic cooperativity with respect to aspartate. In addition, the Asp-100-->Ala mutation restores the ability of the Glu-50-->Ala enzyme to be activated by succinate and increases the affinity of the enzyme for the bisubstrate analogue N-(phosphonacetyl)-L-aspartate (PALA). At subsaturating concentrations of aspartate, the Glu-50/Asp-100-->Ala enzyme is activated more by ATP than the Glu-50-->Ala enzyme and is also inhibited more by CTP than either the wild-type or the Glu-50-->Ala enzyme. As opposed to the wild-type enzyme, the Glu-50/Asp-100-->Ala enzyme is activated by ATP and inhibited by CTP at saturating concentrations of aspartate. Structural analysis of the Glu-50/Asp-100-->Ala enzyme by solution X-ray scattering indicates that the double mutant exists in the same T quaternary structure as the wild-type enzyme in the absence of ligands and in the same R quaternary structure in the presence of saturating PALA. However, saturating concentrations of carbamoyl phosphate and succinate only convert a fraction of the Glu-50/Asp-100-->Ala enzyme population to the R quaternary structure, a behavior intermediate between that observed for the Glu-50-->Ala and wild-type enzymes. Solution X-ray scattering was also used to investigate the structural consequences of nucleotide binding to the Glu-50/Asp-100-->Ala enzyme.

Adenosine Triphosphate↗

PubMed1995

Overexpression and purification of the trimeric aspartate transcarbamoylase from Bacillus subtilis.

A procedure has been developed for the overexpression and purification of milligram quantities of the Bacillus subtilis aspartate transcarbamoylase. The plasmid pEK171, carrying the B. subtilis pyrB structural gene under the control of the Escherichia coli pyrBI promoter, was transformed into the E. coli strain EK1104 and the enzyme overexpressed to approximately 50% of total soluble protein under extreme derepression of the pyrimidine pathway. The enzyme was subsequently purified by means of ammonium sulfate fractionation, anionic exchange chromatography using Q-Sepharose Fast Flow resin, negative chromatography on Matrex Gel Red A agarose, and hydrophobic interaction chromatography using Matrex Phenyl Cellufine. The purification yields approximately 60 mg of pure enzyme per liter of bacterial culture. Kinetic analysis of the overexpressed enzyme indicated that it had kinetic properties very similar to those of the enzyme purified from B. subtilis cells.

Aspartate Carbamoyltransferase↗

PubMed1995

Stereospecificity of hydride removal from NADH by reductases of multicomponent nonheme iron oxygenase systems.

The stereospecificity of hydride removal from the 4 position of the pyridine ring of NADH by reductases from all three classes of multicomponent nonheme iron oxygenases was examined. The class I and II reductases, modules of which show significant sequence similarity with and which belong to the ferredoxin-NADP+ reductase family of flavin-dependent oxidoreductases, transferred the pro-R hydrogen. By contrast, the class II enzymes, which do not show significant sequence similarity to the class I and III enzymes but modules of which belong to the glutathione reductase family of flavoenzymes, transferred the pro-S hydrogen.

Dioxygenases↗

PubMed1995

Evaluation of the Abuscreen ONLINE assay for amphetamines on the Hitachi 737: comparison with EMIT and GC/MS methods.

The performance of the ONLINE Assay for Amphetamines on the Hitachi 737 was compared to the Syva Emit d.a.u. Assay and GC/MS. Randomly screened (n = 2964) patient urine samples were assayed using ONLINE and Emit d.a.u. assays concurrently, using d-amphetamine, 1000 ng/mL and d-methamphetamine, 1000 ng/mL as the screening cutoff for ONLINE and Emit d.a.u. assays, respectively. All presumptive positives were confirmed by GC/MS. The specificity was 99% (2937/2964) for ONLINE and 97% (2873/2964) for Emit. Agreement with GC/MS was 80% (110/137) for ONLINE and 55% (110/201) for Emit.

Amphetamines↗

PubMed1994

Glutamic acid 86 is important for positioning the 80's loop and arginine 54 at the active site of Escherichia coli aspartate transcarbamoylase and for the structural stabilization of the C1-C2 interface.

Glu-86, which interacts with the side chain of Arg-54 across the C1-C2 interface of Escherichia coli aspartate transcarbamoylase, tethers the end of the flexible 80's loop, which moves into the active site during the T to R transition. In order to determine whether this interaction is important for the correct positioning of the 80's loop and Arg-54 at the active site and also for the structural stabilization of the enzyme, a mutant version was created in which Glu-86 was replaced by Gln (Glu-86-->Gln). Although the mutant holoenzyme exhibits almost normal homotropic cooperativity, both the holoenzyme and catalytic subunit exhibit substantial reductions in activity and affinity for aspartate and carbamyl phosphate. Furthermore, the mutant holoenzyme shows a marked decrease in the activation by ATP and by the bisubstrate analog N-(phosphonoacetyl)-L-aspartate, reduced inhibition by CTP, as well as reduced affinities for these ligands. Results from molecular dynamics simulations of the Glu-86-->Gln and Glu-86-->Ala enzymes suggest that the positions of the 80's loop and Arg-54 are significantly perturbed by the introduction of these mutations. Taken together, these results indicate that the interaction between Glu-86 and Arg-54 is important for the formation of the high affinity, high activity form of the enzyme by stabilizing the correct position of the 80's loop and Arg-54 at the active site. Heat inactivation experiments also demonstrated that Glu-86 plays a significant role in the structural stabilization of the C1-C2 interface, since the temperature required for loss of half of the activity of the Glu-86-->Gln catalytic subunit is reduced by 5 degrees C relative to the wild-type catalytic subunit.

Aspartate Carbamoyltransferase↗

PubMed1994

Hyperbaric oxygen therapy following radical vulvectomy: an adjunctive therapy to improve wound healing.

The objective of the study was to evaluate the effect of postoperative hyperbaric oxygen (HBO) therapy on wound breakdown following radical vulvectomy. A prospective, observational study was performed on patients undergoing radical vulvectomy from October 1990 to March 1993 at Scott and White Memorial Hospital, Temple, Texas. HBO therapy was initiated in the postoperative period. Retrospective review of all patients coded as radical vulvectomy from 1987 to 1990 provides for a historical comparison. Eight patients were enrolled in the study. Six patients had radical vulvectomies with lymph node dissections (LND) and two did not. One wound breakdown was observed in the HBO-treated patients. Retrospective review identified 22 patients as having undergone radical vulvectomy with or without LND. Seven of 9 patients with LND and 3 of 13 without LND had wound breakdowns. A significant difference (P = 0.035) was found comparing patients treated with LND and HBO to retrospective controls with LND. Hospitalization was shorter in the HBO-treated patients. This small study showed a reduction in wound breakdown for patients undergoing radical vulvectomy with LND and HBO therapy compared to similar patients not treated with HBO. This observation supports the need for randomized studies examining the efficacy of HBO therapy in this group of patients.

Adolescent↗

PubMed1993

Preliminary crystallographic study of D(-)-mandelate dehydrogenase from Rhodotorula graminis.

NAD+ dependent D(-)-mandelate dehydrogenase from the yeast Rhodotorula graminis strain KGX 39 has been crystallized in three different forms using the hanging drop vapour diffusion method at 15 to 20 degrees C. Type I crystals belong to space group P222(1), P22(1)2(1) or P2(1)2(1)2(1) with a = 100.3 A, b = 117.4 A, c = 80.4 A and are likely to contain a dimer in the crystallographic asymmetric unit. They diffract to dmin = 3.0 A. Type II crystals belong to space group P22(1)2(1) or P2(1)2(1)2(1) with a = 187.8 A, b = 122.9 A, c = 72.1 A and contain probably two dimers in the crystallographic asymmetric unit. They diffract to dmin = 1.8 A. Type III crystals belong to space group P2(1)2(1)2(1) with a = 109.6, b = 52.0 A, c = 145.7 A, and are likely to contain a dimer in the crystallographic asymmetric unit. They diffract at least to dmin = 2.5 A.

Alcohol Oxidoreductases↗

PubMed1993

The conserved residues glutamate-37, aspartate-100, and arginine-269 are important for the structural stabilization of Escherichia coli aspartate transcarbamoylase.

Aspartate transcarbamoylase from Escherichia coli is a dodecameric enzyme consisting of two trimeric catalytic subunits and three dimeric regulatory subunits. The X-ray structure of this enzyme indicates that the side chains of His-41, Asp-100, and Asp-90 from one catalytic chain form interactions with the side chains of Glu-37, Arg-65, and Arg-269, respectively, from an adjacent catalytic chain. In order to determine whether these interactions are important for the structural stabilization of the enzyme and/or homotropic and heterotropic effects, four mutant versions of aspartate transcarbamoylase, Glu-37-->Ala, Asp-100-->Asn, Asp-100-->Ala, and Arg-269-->Ala, were created by site-specific mutagenesis. The Glu-37-->Ala holoenzyme exhibits essentially wild-type behavior with respect to homotropic cooperativity and heterotropic regulation by ATP and CTP. The Glu-37-->Ala catalytic subunit exhibits a half-life of inactivation at 69 +/- 0.5 degrees C of 4.9 min, as compared to 5.8 min for the wild-type catalytic subunit. The Asp-100-->Asn and Asp-100-->Ala holoenzymes are slightly more active than the wild-type holoenzyme, exhibit 1.4-fold and 1.8-fold reductions in the aspartate concentration at half the maximal specific activity, respectively, and show increased affinities for ATP and CTP. Both the Asp-100-->Asn and Asp-100--> Ala catalytic subunits exhibit a 2-fold reduction in the half-life of inactivation at 69 +/- 0.5 degrees C.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

PubMed1993

Relationships amongst some bacterial and yeast lactate and mandelate dehydrogenases.

Five yeast strains were isolated by enrichment culture on the basis of their ability to grow on mandelate and two of these strains were identified as Rhodotorula glutinis. In addition, a range of yeasts from culture collections was screened for growth on mandelate. The results suggest that mandelate utilization is a widespread but not universal characteristic within the genus Rhodotorula. Several of the yeasts contained an inducible NAD-dependent D(-)-mandelate dehydrogenase and an inducible dye-linked (presumably flavoprotein) L(+)-mandelate dehydrogenase. All the D(-)-mandelate dehydrogenases from the yeasts showed immunological cross-reactivity with each other (as judged by both immunoinhibition and immunoblotting), as did all the yeast L(+)-mandelate dehydrogenases that were tested. Determination of N-terminal amino acid sequences of several bacterial and yeast lactate and mandelate dehydrogenases, together with the evidence from the immunological studies, confirmed and extended previous proposals that there are several major groups of such dehydrogenases: FMN-dependent, membrane-bound L(+)-lactate and L(+)-mandelate dehydrogenases (M(r) = approx. 44,000) in bacteria, mitochondrial flavocytochrome b2 L(+)-lactate and L(+)-mandelate dehydrogenases (M(r) = approx. 59,000) in yeasts, FAD-dependent, membrane-bound D(-)-lactate and D(-)-mandelate dehydrogenases in bacteria, and soluble NAD-dependent D(-)-mandelate dehydrogenases in both bacteria and yeasts.

Alcohol Oxidoreductases↗

PubMed1992

Mechanistic and active-site studies on D(--)-mandelate dehydrogenase from Rhodotorula graminis.

D(--)-Mandelate dehydrogenase, the first enzyme of the mandelate pathway in the yeast Rhodotorula graminis, catalyses the NAD(+)-dependent oxidation of D(--)-mandelate to phenylglyoxylate. D(--)-2-(Bromoethanoyloxy)-2-phenylethanoic acid ['D(--)-bromoacetylmandelic acid'], an analogue of the natural substrate, was synthesized as a probe for reactive and accessible nucleophilic groups within the active site of the enzyme. D(--)-Mandelate dehydrogenase was inactivated by D(--)-bromoacetylmandelate in a psuedo-first-order process. D(--)-Mandelate protected against inactivation, suggesting that the residue that reacts with the inhibitor is located at or near the active site. Complete inactivation of the enzyme resulted in the incorporation of approx. 1 mol of label/mol of enzyme subunit. D(--)-Mandelate dehydrogenase that had been inactivated with 14C-labelled D(--)-bromoacetylmandelate was digested with trypsin; there was substantial incorporation of 14C into two tryptic-digest peptides, and this was lowered in the presence of substrate. One of the tryptic peptides had the sequence Val-Xaa-Leu-Glu-Ile-Gly-Lys, with the residue at the second position being the site of radiolabel incorporation. The complete sequence of the second peptide was not determined, but it was probably an N-terminally extended version of the first peptide. High-voltage electrophoresis of the products of hydrolysis of modified protein showed that the major peak of radioactivity co-migrated with N tau-carboxymethylhistidine, indicating that a histidine residue at the active site of the enzyme is the most likely nucleophile with which D(--)-bromoacetylmandelate reacts. D(--)-Mandelate dehydrogenase was incubated with phenylglyoxylate and either (4S)-[4-3H]NADH or (4R)-[4-3H]NADH and then the resulting D(--)-mandelate and NAD+ were isolated. The enzyme transferred the pro-R-hydrogen atom from NADH during the reduction of phenylglyoxylate. The results are discussed with particular reference to the possibility that this enzyme evolved by the recruitment of a 2-hydroxy acid dehydrogenase from another metabolic pathway.

Affinity Labels↗

PubMed1990

Estrogen-replacement therapy in patients with previous endometrial carcinoma.

For patients with previous endometrial cancer, ERT is not the accepted practice in the U.S. The therapeutic dictum that estrogen is contraindicated in patients with previous uterine adenocarcinoma is, however, not substantiated by clinical data. The relation of unopposed estrogen stimulation to endometrial hyperplasia and carcinoma, and the published studies relating ERT to endometrial cancer, have resulted in the clinical perception--and cautionary statements to that effect--that estrogen is contraindicated for patients with a history of endometrial carcinoma. The exact biologic effects of ERT on endometrial adenocarcinoma have not yet been studied adequately, however; the initial clinical data suggest that there is no increase in recurrence or mortality. In the meantime, the clinician is left with contradictory data as a basis for determining the proper management of symptomatic patients. The total impact of estrogen deficiency on the health of women and the ratio of benefits and risks of ERT are yet to be defined completely. The preponderance of evidence suggests that estrogen has a beneficial effect on the major cause of death in women, coronary heart disease, by increasing the high-density lipoprotein (HDL) fraction of cholesterol. It is established that estrogen prevents the demineralization of bone and delays the ravages of osteoporosis. No one has died from vaginal atrophy, bladder dysfunction, or hot flashes; the quality of life and marriage have been improved, however, by relieving these symptomatic conditions with ERT. Several studies have attempted to analyze with various statistical models the ratio of benefits to risks, and the majority of authors have concluded that the beneficial effect on cardiovascular disease alone clearly outweighs any known risk.(ABSTRACT TRUNCATED AT 250 WORDS)

Adenocarcinoma↗