Survival after elective surgery for colonic cancer in Denmark.

AIM: Total mesorectal excision (TME) has been shown to improve the outcome for patients with rectal cancer. In contrast, there are fewer data on complete mesocolic excision (CME) for colonic cancer. METHOD: Data from the National Colorectal Cancer Database were analysed. This includes about 95% of all patients with colorectal cancer in Denmark. Only patients having elective surgery for colonic cancer in the period 2001-2008 were included. Overall and relative survival analyses were carried out. The study period was divided into the periods 2001-2004 and 2005-2008. RESULTS: 9149 patients were included for the final analysis. The overall 5-year survival rates were 0.65 in 2001-2004 and 0.66 in 2005-2008. The relative 5-year survival rates were also within 1% of each other. None of these comparisons was statistically significant. CONCLUSION: Survival following elective colon cancer surgery has been almost unchanged since 2001.

Epidemiology of erectile dysfunction and its risk factors: a practice-based study in Denmark.

The objective of this study was to estimate the prevalence of erectile dysfunction (ED) and its health-related correlates among Danish men, to evaluate the influence of age, tobacco smoking, educational level and medication and the needs for treatment and willingness to be treated. A validated questionnaire was sent to 4310 noninstitutionalized Danish men, aged 40-80 y. The men selected constituted all male patients aged 40-80 y in 12 general practitioner practices in a county of Zealand, representing both the urban and rural population. Besides age, education, marital status and International Index of Erectile Function, the questionnaire included the duration of sexual problems (ED, premature ejaculation, penile curvature), comorbidity, medication, risk factors and the effect of prior treatment and willingness to seek treatment for sexual problems. A total of 2210 men responded, giving a response rate of 51.3%. No difference in the response rate by age groups was noted. The prevalence of complete ED increased with increasing age: 40-45 y, ED: 4.5%; 50-55 y, ED: 11.1%; and 75-80 y ED: 52%. The frequency of ED increased three-fold from men without comedication to men having some kind of medical treatment. Risk factors included tobacco smoking and low educational level. Only 9% suffering from ED had received some kind of treatment. Of the treated men, 75% were satisfied with the treatment. Willingness to discuss sexual matters depended both upon the age of the man and his actual erectile function. Taboos were seen more frequently among elderly people. ED increases with age, but only 10% of the men with sexual problems seek advice. Medication predisposes to ED.

Dynamics of mononuclear cadmium beta-lactamase revealed by the combination of NMR and PAC spectroscopy.

The two metal sites in cadmium substituted beta-lactamase from Bacillus cereus 569/H/9 have been studied by NMR spectroscopy ((1)H, (15)N, and (113)Cd) and PAC spectroscopy ((111m)Cd). Distinct NMR signals from the backbone amides are identified for the apoenzyme and the mononuclear and binuclear cadmium enzymes. For the binuclear cadmium enzyme, two (113)Cd NMR signals (142 and 262 ppm) and two (111m)Cd PAC nuclear quadrupole interactions are observed. Two nuclear quadrupole interactions are also observed, with approximately equal occupancy, in the PAC spectra at cadmium/enzyme ratios < 1; these are different from those derived for the binuclear cadmium enzyme, demonstrating interaction between the two metal ion binding sites. In contrast to the observation from PAC spectroscopy, only one (113)Cd NMR signal (176 ppm) is observed at cadmium/enzyme ratios < 1. The titration of the metal site imidazole (N)H proton signals as a function of cadmium ion-to-enzyme ratio shows that signals characteristic for the binuclear cadmium enzyme appear when the cadmium ion-to-enzyme ratio is between 1 and 2, whereas no signals are observed at stoichiometries less than 1. The simplest explanation consistent with all data is that, at cadmium/enzyme ratios < 1, the single Cd(II) is undergoing exchange between the two metal sites on the enzyme. This exchange must be fast on the (113)Cd NMR time scale and slow on the (111m)Cd PAC time scale and must thus occur in a time regime between 0.1 and 10 micros.

[Rehabilitation of patients with chronic obstructive lung disease. Are two exercise sessions per week sufficient?]. / Rehabilitering af patienter med kronisk obstruktiv lungesygdom. Er troening to gange ugentlig tilstroekkeligt?

Several studies of patients with chronic obstructive pulmonary disease (COPD) have shown that pulmonary rehabilitation three to seven times a week improves exercise performance and well being. This study investigates feasibility, effect and economic aspects of a programme consisting of two sessions a week. Twenty-four patients were randomized to rehabilitation and twenty-one to placebo. In an outpatient setting patients were assigned to an eight-week programme of exercise plus education twice a week (Exercise group) or conventional community cares (Placebo group). Seven patients did not complete the rehabilitation. The characteristics of the thirty-eight COPD patients at baseline (mean +/- SD): forced expiratory volume in one second (FEV-1) 1.1 +/- 0.4 L, six-minute walk distance (6MWD) 413 +/- 75 m. Rehabilitation resulted in an insignificant improvement in well being and the 6MWD (29 m ¿95% confidence interval: -8-66 m¿. Rehabilitation session twice a week for eight weeks had no effect in patients with moderate COPD.

Rehabilitation of patients with chronic obstructive pulmonary disease. Exercise twice a week is not sufficient!

Several studies of chronic obstructive pulmonary disease (COPD) have shown that pulmonary rehabilitation, consisting of at least three training sessions a week, improves exercise performance and health status. This study investigates feasibility, effect and economic aspects of a rehabilitation programme consisting of two sessions a week for 8 weeks. Twenty-four patients with moderate COPD were randomized to rehabilitation and 21 to placebo. Patients were assigned to an 8-week programme of exercise plus education (Exercise group) or conventional community care (Placebo group). The rehabilitation program was carried out in a hospital outpatient setting and consisted of 16 h exercise and 13.5 h of education. The exercise group received physiotherapy and education twice a week. Seven patients did not complete the programme. The characteristics of the 38 COPD-patients at baseline were the following: (mean +/- SD) forced expiratory volume in 1 sec (FEV1) 1.1+/-0.4 1 (47% of predicted), 6-min walking distance (6MWD) 413+/-75 m, score of St. George's Respiratory Questionnaire (SGRQ) 44+/-21. Health-status, assessed by SGRQ and The Psychological General Well-being (PGWB) Index, did not improve. Rehabilitation resulted in an insignificant improvement in the 6MWD [29 m (95% confidence interval: -8 -66 m)]. We conclude that a rehabilitation program consisting of exercise and education twice a week for 8 weeks had no effect on exercise performance and well being in patients with moderate COPD.

Interaction and coordination geometries for Ag(I) in the two metal sites of hemocyanin.

111Ag(I) perturbed angular correlations of gamma-rays (PAC) has been used to investigate the binuclear metal site of 111Ag(I)-substituted Carcinus aestuarii deoxyhemocyanin. The studies have shown that apo-hemocyanin is able to bind 2 mol of Ag(I) per mol of protein and that the binding is specific for the metal ion sites. The PAC spectra show pronounced changes when the stoichiometry of Ag(I) to protein is increased from 0.1 to 2.0. These changes have been interpreted as evidence of interactions between the two sites in terms of a structural destabilization of the first occupied site caused by the occupation of the second site. The experimental data for the Ag(I)-substituted metal sites do not agree well with the three-coordinated structure found in the Cu(I) holo-protein. However, if a water molecule is included as a coordinating ligand in the Ag(I) metal site a successful interpretation of the experimental data can be obtained.

Preference of Cd(II) and Zn(II) for the two metal sites in Bacillus cereus beta-lactamase II: A perturbed angular correlation of gamma-rays spectroscopic study.

Cd-substituted forms of the Bacillus cereus metallo-beta-lactamases (BCII) were studied by perturbed angular correlation of gamma-rays (PAC) spectroscopy. At very low [Cd]:[apo-beta-lactamase] ratios, two nuclear quadrupole interactions (NQI) were detected. For [Cd]:[apo-beta-lactamase] ratios between 0.8 and 3.0, two new NQIs appear, and the spectra show that up to 2 cadmium ions can be bound per molecule of apoenzyme. These results show the existence of two interacting Cd-binding sites in BCII. The relative populations of the two NQIs found at low [Cd]:[apo-beta-lactamase] ratios yielded a 1:3 ratio for the microscopic dissociation constants of the two different metal sites (when only one cadmium ion is bound). X-ray diffraction data at pH 7.5 demonstrate that also for Zn(II) two binding sites exist, which may be bridged by a solvent molecule. The measured NQIs could be assigned to the site with three histidines as metal ligands (three-His site) and to the site with histidine, cysteine, and aspartic acid as metal ligands (Cys site), respectively, by PAC measurements on the Cys168Ala mutant enzyme. This assignment shows that cadmium ions preferentially bind to the Cys site. This is in contrast to the preference of Zn(II) in the hybrid Zn(II)Cd(II) enzyme, where an analysis of the corresponding PAC spectrum showed that Cd(II) occupied the Cys site, whereby Zn(II) occupied the site with three histidines. The difference between Zn(II) and Cd(II) in affinity for the two sites is combined with the kinetics of hydrolysis of nitrocefin for different metal ion substitutions (Zn(2)E, ZnE, Cd(2)E, CdE, and ZnCdE) to study the function of the two metal ion binding sites.

Plastocyanin binding to photosystem I as a function of the charge state of the metal ion: effect of metal site conformation.

The binding of Ag- and Cd-substituted plastocyanin to reduced photosystem 1 of spinach has been studied through the rotational correlation time of plastocyanin measured by the technique of perturbed angular correlation of gamma-rays (PAC). Ag and Cd are used as models for native Cu(I) and Cu(II), respectively. A dissociation constant of 5 microM was found for Ag-plastocyanin, whereas the dissociation constant was at least 24 times higher for Cd-plastocyanin. PAC was further used to characterize the structure of the metal site of Cd- and Ag-plastocyanin. The Cd spectra are characteristic of a planar configuration of one cysteine and two histidines. However, the spectra show an unusual peak broadening and a high degree of internal motion, interpreted as motion of one of the histidines within the plane. (111)Ag decays to (111)Cd, followed by the emission of two gamma-rays used for the PAC experiment. The (111)Ag PAC spectra indicate that one of the coordinating histidines has a different position in the Ag protein than in the Cd protein but that the decay of Ag to Cd causes a relaxation of the position of this histidine to the position in the Cd protein within 20 ns. Binding of Ag-plastocyanin to photosystem I stabilized the Ag metal site structure so that no relaxation was observed on a time scale of 100 ns. This stabilization of the Ag structure upon binding indicates that the metal site structure is involved in regulating how the dissociation constant for plastocyanin depends on the charge of the metal ion.

Structure, Chemical Bonding, and Nuclear Quadrupole Interactions of beta-Cd(OH)(2): Experiment and First Principles Calculations.

The structure of Cd(OH)(2) was determined by X-ray diffraction on powder crystals and by calculations using the full-potential linearized augmented plane wave method. Good agreement between the two results was found. The chemical bonding is characterized by the interactions of the OH(-) group with Cd(2+) which is not only electrostatic but shows some polarization or covalent admixtures and by the covalent bond in the OH(-) group. The electric field gradient (EFG) was calculated and compared with an experimental determination of the nuclear quadrupole interaction using perturbed angular correlation of gamma-rays. The calculated EFG agrees well with the EFG derived from experiment. The total electric field gradient was decomposed into contributions from different orbitals and energy regions showing that both the Cd 5p and 4d wave functions contribute significantly. Finally, the influence of spin-orbit coupling on the electric field gradient was investigated and found to be of little importance.

Structure and dynamics of the metal site of cadmium-substituted carboxypeptidase A in solution and crystalline states and under steady-state peptide hydrolysis.

PAC spectra (perturbed angular correlation of gamma-rays) of cadmium-substituted carboxypeptidase A (CPD) show that the enzyme in solution imposes a flexible, pH- and chloride-dependent coordination structure on the metal site, in contrast to what is found in the crystalline state. A much more restricted coordination geometry occurs for the steady-state peptide intermediates of Bz-Gly-l-Phe and Bz-Gly-Gly-l-Phe in solution, suggesting that substrate binding locks the structure in a rigid conformation. The results further indicate that the peptide intermediate has a six-coordinated metal coordination geometry with an OH- ligand at the solvent site and a carbonyl oxygen at an additional ligand site. In marked contrast, conformational rigidity is not induced by the inhibitor/poor substrate Gly-L-Tyr nor by the products of high turnover substrates, Bz-Gly, Bz-Gly-Gly, and L-Phe. These results are consistent with an intact scissile peptide bond in the enzyme-substrate complex of Bz-Gly-L-Phe and Bz-Gly-Gly-L-Phe. A single nuclear quadrupole interaction (NQI) is observed for the crystalline state of the enzyme between pH 5.7 and pH 9.4. This NQI agrees with calculations based on the metal coordination geometry for cadmium in crystalline CPD derived from X-ray diffraction studies. A single broad distribution of NQIs is observed for CPD in sucrose solutions and 0.1 M NaCl at pH values below 6.5. This NQI (NQI-1') has parameters very close to those for the crystalline state. The enzyme metal site, characterized by this NQI, is converted into two new enzyme metal sites over the pH range of 6.5-8.3. The metal coordination sphere of one of these has a NQI (NQI-1) with parameters similar to those at lower pH values (NQI-1') while the other NQI (NQI-2) is characterized by markedly different NQI parameters. Angular overlap model (AOM) calculations indicate that the coordination sites giving NQI-1' and NQI-1 both have a metal-bound water molecule while the coordination site giving NQI-2 has a metal-bound hydroxide ion. PAC results at pH 8.3-10.5 indicate that in this pH range the two metal coordination geometries related to NQI-1 and NQI-2 occur in a pH independent ratio of 2:1, with the one with the water ligand being the most abundant species. The observed pH-independent equilibrium between the two different metal coordination geometries for cadmium can be explained by an equilibrium between tautomeric forms of a hydrogen bond between the Glu-270 carboxyl group and the metal-bound water (Glu-270 COO-...(HOH)M <==> Glu-270 COOH...(OH-)M) being slow on the time scale of a PAC experiment, i.e., slower than 0.5 micros. We finally suggest that NQI-1' observed at low pH reflects an enzyme species containing a metal-coordinated water molecule and the protonated carboxyl group of Glu-270.

Coordination geometries for monovalent and divalent metal ions in [His121]azurin--studies using perturbed angular correlations of gamma-rays from 111Ag and 111mCd.

The structural details of the metal site in the [His121]azurin mutant from Alcaligenes denitrificans where the axial methionine has been replaced by a histidine have been studied after substitution with the divalent cadmium ion and the monovalent silver ion. The studies have been carried out in solution using the technique of perturbed angular correlations of gamma-rays (PAC) of the two isotopes, 111Ag and 111mCd. In the pH range 6-9, the PAC spectra for cadmium-substituted [His121]azurin reveals a pH-independent equilibrium between two different metal-coordination geometries. Interpretation of the PAC data shows agreement between the dominating coordination geometry and that derived from X-ray diffraction on the Cu(II)[His121] azurin at high pH (Messerschmidt, A., unpublished results). Thus, it appears likely that cadmium for this geometry is four coordinated to the ligands His46, His117, Cys112, and His121. The other geometry is best interpreted as a substitution of His121 by a solvent water ligand. These interpretations stem from predictions of the experimentally determined nuclear quadrupole interactions (NQI) via the simple angular overlap model (AOM). At low pH (3.8), the concentration of the former species is reduced to 50% of its high pH value suggesting a pK of about 4 for His121. Two different coordination geometries have also been observed for the Cu(II) protein and assigned a type 1.5 and a type 1 copper site [Kroes, S. J., Hoitink, C. W. G., Andrew, C. R., Ai, J., Sanders-Loehr, J., Messerschmidt, A., Hagen, W. R. & Canters, G. W. (1996a) Eur. J. Biochem. 240, 342-351]. For silver-substituted [His121]azurin, several notable changes occur relative to the cadmium-substituted protein. At least four different metal-coordination geometries exist for silver[His121]azurin in the pH range 4-8. Changes in the population of these coordination sites occurs between pH 4 and pH 5, and pH 5 and pH 6. Furthermore, in contrast to the cadmium-substituted protein, only a single coordination geometry is present above pH 6. The change in population occurring between pH 5 and pH 6 suggests ionization of a non-coordinating histidine, here proposed as His121. The change in population at low pH could then be due to protonation of an additional coordinating histidine such as His46 or His117. The single coordination geometry existing at pH values above 6 for the silver protein cannot within our model calculations be described with His121 coordinated. However, it can be described with a coordinated water molecule but in a different angular position than for His121 in the copper protein (Messerschmidt, A., unpublished results). The reduced tendency for silver to coordinate His121 is in agreement with the general trend of lower pK values for ligands coordinating to monovalent ions relative to divalent ions. In conclusion, this work demonstrates that mutation of Met121 to other amino acid residues opens the possibility of other coordination geometries than the rigid three-coordinated structure observed for wild-type azurin, especially the possibility of increasing the coordination number by either solvent water ligands or the substituting amino acid. Furthermore, it opens up the possibility for different coordination geometries for monovalent and divalent metal ions as observed here and previously for the [Leu121]azurin mutant [Bauer, R., Danielsen, E., Hemmingsen, L., Bjerrum, M. J., Hansson, O. & Singh, K. (1997) J. Am. Chem. Soc. 119, 157-163].

The protein conformation of Cd-substituted horse liver alcohol dehydrogenase and its metal-site coordination geometry in binary and ternary inhibitor complexes.

The coordination geometry of the metal at the active site in Cd-substituted horse liver alcohol dehydrogenase (LADH) has been investigated for the binary complexes of LADH with imidazole, isobutyramide, decanoic acid and Cl-, and for the ternary complexes of LADH with NADH and imidazole, NADH and isobutyramide, NAD+ and decanoic acid and NAD+ and Cl-, by using the method of perturbed angular correlation of gamma-rays (PAC). The spectral results are consistent with a flexible structure around the metal for the binary complexes with inhibitors. For ternary complexes, however, a rigid structure is observed. An exception is the ternary complex between LADH, NADH and imidazole, in which the metal site is still flexible. Comparing with available structures determined by X-ray crystallography, we found a correlation between open structures and flexible metal sites, and between closed structures and rigid metal sites. This indicates that the PAC technique can be applied to distinguish the two conformations in solution. The spectral parameters, omega(o) and eta, of the experiments, except for the complexes with imidazole, fall into two groups: one with low omega(o) and one with high omega(o) (eta is relatively constant in all experiments). In this work it is clarified that the low omega(o) values are connected with the presence of a negatively charged solvent ligand. Using an angular-overlap approach to interpret the results, the low omega(o) values are found to be compatible with a coordination geometry where the S-Cd-S (Cys174 and Cys46 coordinate to the metal) angle is about 110 degrees as suggested in [Hemmingsen, L., Bauer, R., Danielsen, E., Bjerrum. M. J., Zeppezauer, M., Adolph, H. W., Formicka, G. & Cedergren-Zeppezauer, E. (1995) Biochemistry 34, 7145-7153], whereas high omega(o) values are compatible with an S-Cd-S angle of 130 degrees. The presence of a negatively charged metal ligand, therefore, might trigger the movement of the sulfur of Cys174. As it is believed that alcohols coordinate to the metal as alcoholate ions this could be important for catalysis.

alpha1-acid glycoprotein and albumin in the spinal fluid of pregnant women.

Pregnant women at term need a smaller dose of local anaesthetic to produce the same spinal anaesthesia as non-pregnant women. The aim of the study was to see whether the concentrations of binding proteins were reduced in spinal fluid in pregnant women. The concentrations of the binding proteins [Formula: see text] glycoprotein (AAG) and albumin were measured in plasma and spinal fluid from 20 pregnant and 20 non-pregnant women, ASA 1. Mean spinal fluid albumin was 3.13 micromol/1 in non-pregnant and 1.97 micromol/1 in pregnant women (95% confidence interval of the difference: 0.62 - 1.50). Mean spinal fluid AAG was 0.152 micromol/1 in non-pregnant and 0.081 micromol/1 in pregnant women (95% confidence interval of the difference: 0.030 - 0.100). The concentrations in plasma were also reduced in late pregnancy. All the changes were statistically significant. Since only a minor fraction of bupivacaine seems to be protein bound in spinal fluid these differences may have only a little influence on the free fraction and cannot be solely responsible for the increased sensitivity of pregnant women to spinal anaesthesia.

Effect of inhibitors on the coordination geometries of cadmium at the metal sites in angiotensin-I-converting enzyme.

Perturbed angular correlation of gamma-rays (PAC) spectroscopy has been used to investigate the angiotensin-I-converting enzyme (ACE) of rabbit lung. By substituting the zinc ions in ACE with excited 111mCd2+ ions, analysis of PAC spectra gave directly the percentage of cadmium ions bound to ACE. The result of the analysis was a dissociation constant of about 1 microM for the cadmium-ACE complex, and a stoichiometry of two moles cadmium/mole enzyme. Cadmium binding is thus about two orders of magnitude weaker than zinc binding to ACE but two orders of magnitude stronger than cobalt binding. PAC spectra monitor the nuclear quadrupole interaction (NQI) for 111mCd. The NQI for ACE exhibits very low frequencies in the PAC spectra with a rather large spectral broadening. In the presence of the inhibitor ramiprilat, the frequencies increase but the spectral broadening is about the same as for ACE without inhibitor. When the inhibitor captopril is added, very high frequencies are obtained consistent with sulfur binding, but now with a narrower distribution of NQI's. A simple molecular orbital analysis of the obtained NQI's has been performed, using a coordination sphere of two His, one Glu residue and a solvent ligand, equivalent to the zinc ligands in thermolysin and carboxypeptidase. The calculated spectral parameters could be modelled with the measured parameters if the solvent ligand is H2O in free ACE, carboxylate from ramiprilat in the ACE-ramiprilat complex and a mercapto group in the ACE-captopril complex. The coordination geometry for cadmium carboxypeptidase obtained by X-ray diffraction gives a calculated set of NQI parameters consistent with the measured parameters for cadmium in the captopril-ACE complex using a mercapto group as the solvent ligand. However, for ACE and its complex with ramiprilat, a significant distortion of the cadmium geometry for carboxypeptidase A had to be adopted in order to calculate NQI's close to the experimental values.

Structure of metal site in Cd-substituted His117Gly mutant of azurin with and without addition of imidazole derivatives.

The present work uses 111mCd-perturbed angular correlations of gamma-rays (PAC) to investigate the structure of the metal site of the His117Gly mutant of Pseudomonas aeruginosa azurin in aqueous solution and the effect on the structure upon addition of the following exogenous ligands: imidazole, 4-methyl imidazole, 1-methyl imidazole, 2-methyl imidazole and histidine. The nuclear quadrupole interaction of cadmium bound to the mutant without addition of exogenous ligands shows a strong pH dependence with three different nuclear quadrupole interactions consistent with two pKa values at about 7.2 and 8.6 at 2 degrees C. Addition of the imidazole derivatives resulted in a significant change in the PAC spectrum showing that they coordinate. This is in accordance with observations by EPR for the same mutant with copper at the metal site [den Blaauwen, T. & Canters, G. W. (1993) J. Am. Chem. Soc. 115, 1121-1129]. However, whereas EPR and ultraviolet/visual absorption show that the characteristics of the wild-type copper protein are regained by addition of the imidazole derivatives with the exception of the possible bidentates (histidine and histamine), the comparison of the PAC results to model calculations shows that the cadmium ion must be fourfold coordinated in most cases, probably binding an additional water or hydroxide ligand. A fourfold coordination is in contrast to cadmium-substituted wild-type azurin where PAC data inferred a threefold coordination by a Cys and two His residues [Danielsen, E. Bauer, R., Hemmingsen, L., Andersen. M., Bjerrum, M. J., Butz, T., Tröger, W., Canters, G. W., Hoitink, C. W. G., Karlsson, G., Hansson, O. & Messerschmidt, A. (1995) J. Biol. Chem. 270, 573-580]

Urine profiles and kidney histology after ionic and nonionic radiologic and magnetic resonance contrast media in rats with cisplatin nephropathy.

RATIONALE AND OBJECTIVES: The nephrotoxic drug cisplatin has been used successfully in treating some cancers. Patients with suspected carcinoma frequently undergo examinations with contrast media. We examined whether ionic and nonionic radiologic and magnetic resonance contrast media would have any effect on cisplatin nephropathy in rats. METHODS: Urine and serum profiles were monitored for 24 days after intravenous (i.v.) injections of saline, diatrizoate, iohexol, gadopentetate dimeglumine, and gadodiamide in high doses (4.59 mmol/kg body weight) in rats that received a weekly intraperitoneal (i.p.) injection of cisplatin (1 mg/kg) for 10 weeks. There were 10 rats in each group. Another 10 rats injected with both i.p. and i.v. saline served as control subjects. After euthanization, rats' kidneys were removed for examination by light microscopy and electron microscopy. RESULTS: Light and electron microscopy showed severe morphologic changes, including tubular dilatation, atrophy, and necrosis induced by cisplatin; however, the contrast media did not induce any additional morphologic changes. Gadopentetate dimeglumine, diatrizoate, and iohexol significantly increased (3-20 times) albuminuria compared with i.v. saline in cisplatin nephropathy, whereas gadodiamide did not. Albuminuria was highest after diatrizoate injection. All four contrast media caused an immediate and transient significant increase in the excretion of the brush border enzymes alkaline phosphatase and gamma-glutamyltransferase (125-500 times) and the cytoplasmatic enzymes alanine aminopeptidase and lactate dehydrogenase (16-100 times). Compared with saline, the ionic agents significantly increased the excretion of both glucose (two times) and sodium (three to five times), whereas the nonionic agents did not. CONCLUSION: High doses of radiologic and magnetic resonance contrast agents cause temporary dysfunction in rats with cisplatin nephropathy. Gadodiamide caused the least dysfunction and diatrizoate the most.

Cd-substituted horse liver alcohol dehydrogenase: catalytic site metal coordination geometry and protein conformation.

The coordination geometry of the catalytic site in Cd-substituted horse liver alcohol dehydrogenase (LADH) has been investigated as a function of pH using the method of perturbed angular correlation of gamma-rays (PAC). LADH in solution fully loaded with cadmium, including radioactive 111mCd in the catalytic site [Cd2(111mCd)Cd2LADH], was studied over the pH range 7.9-11.5. Analysis of the PAC spectra showed the ionization of a group with pKa of 11. This pKa value is about 2 pH units higher than that of native zinc-containing LADH. A pKa of 9.6 was found for the binary complex of Cd2(111mCd)Cd2LADH with NAD+. This value is also about 2 pH units higher than that of the binary complex of native zinc-containing enzyme and NAD+. No pH dependency was detected for the binary complex of Cd2(111mCd)Cd2LADH with NADH within the pH range measured (pH 8.3-11.5). Assuming that metal-coordinated water is the ionizing group [Kvassman, J., & Pettersson, G. (1979) Eur. J. Biochem. 100, 115-123], we conclude that the larger ionic radius of Cd(II) relative to Zn(II) in the catalytic site causes the elevated pKa values of metal-bound water. Interpretation of nuclear quadrupole interaction (NQI) parameters derived from PAC spectra is based on the use of the angular overlap model, using the coordinates for the catalytic zinc site from the 1.8 A resolution crystal structure of the ternary complex between LADH, NADH, and dimethyl sulfoxide as a model.(ABSTRACT TRUNCATED AT 250 WORDS)

Structure of metal site in azurin, Met121 mutants of azurin, and stellacyanin investigated by 111mCd perturbed angular correlation (PAC).

The geometries of the metal sites in cadmium-substituted azurins have been investigated by 111mCd perturbed angular correlation (PAC). The study includes wild type azurin as well as Met121 mutants of azurin, where methionine has been substituted by Ala, Asn, Asp, Gln, Glu, and Leu. The nuclear quadrupole interaction of wild type azurin analyzed in the angular overlap model is well described as coordination of His46, His117, and Cys112 and cannot be described by coordination of Met121 and/or Gly45. For most of the mutants, there exist two coordination geometries of the cadmium ion. With the exception of the Glu and Asp mutants, one of the conformations is similar to the wild type conformation. The other coordination geometries are either best described by a coordinating water molecule close to the original methionine position or by coordination by the substituting amino acid. These experiments show that even though the methionine does not coordinate it plays an important role for the geometry of the metal site. The nuclear quadrupole interaction of stellacyanin was also measured. The value resembles the most prominent nuclear quadrupole interaction of the Met121-->Gln mutant of Alcaligenes denitrificans azurin, indicating that the structures of the two metal sites are similar.