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L Hemmingsen

Publications and source records attributed to L Hemmingsen.

At least 19 recordsLinked to original sources
PubMed1995

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.

Angiotensin-Converting Enzyme Inhibitors↗
PubMed1995

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]

Azurin↗
PubMed1995

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)

Alcohol Dehydrogenase↗
PubMed1995

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.

Azurin↗
PubMed1994

Retinopathy in type II diabetes mellitus associated with above-normal urinary excretion of RBP.

We performed a cross-sectional study on the urinary excretion profiles of albumin (a marker of glomerular dysfunction) and retinol-binding protein (a low molecular mass protein marker of renal proximal tubular dysfunction) in non-insulin dependent (Type II) diabetics, with or without retinopathy. The urinary excretion of both proteins, in particular retinol-binding protein, was significantly higher in patients with background/proliferative retinopathy compared to patients without retinopathy. The degree of retinopathy correlated to the urinary excretion of albumin (P < 0.005) and retinol-binding protein (P < 0.0001). Retinopathy occurred at a higher frequency in patients with above-normal urinary excretion of retinol-binding protein, both in the absence or presence of micro/macroalbuminuria. The frequency of retinopathy among micro/macroalbuminuric patients with a normal urinary excretion of retinol-binding protein did not differ significantly from that observed in patients with a normal urinary excretion of both proteins. We cannot explain the association between retinopathy and proximal tubular dysfunction in Type II diabetes. However, it is possible that both phenomena are related to a common pathogenetic factor.

Aged↗
PubMed1993

Urine profiles and kidney histologic findings after intravenous injection of mannitol and iohexol in the degeneration phase of gentamicin nephropathy in rats.

RATIONALE AND OBJECTIVES: Previous studies have shown that iodinated contrast media may cause further renal dysfunction in tubulointerstitial nephropathy induced by gentamicin. The current investigation was undertaken to study whether the dysfunction after intravenous injection of a low-osmolar contrast medium is due to a chemotoxic and/or an osmotic effect. METHODS: Urine profiles were followed for 3 or 9 days after intravenous injection of saline, mannitol, and varying dosages of iohexol (1, 2.5, 5, and 10 mL/kg body weight (BW); 350 mg I/mL) in 60 rats, in which intramuscular injection of 40 mg/kg BW gentamicin had been administered daily nine times. A seventh group of 10 rats was given 20 mg/kg BW gentamicin and 5 mL/kg BW of 350 mg I/mL iohexol. Another 10 rats injected with saline served as controls. RESULTS: Both mannitol and iohexol increased the excretion of albumin and the enzyme N-acetyl-B-D-glucosaminidase (NAG) temporarily; the effect was independent of the dose of iohexol. There was a dose-dependent effect on the transient increase in excretion of the enzymes lactate dehydrogenase (LDH), gamma glutamyltransferase (GGT), and alkaline phosphatase (ALK); mannitol did not increase the excretion of these enzymes. In the group given 20 mg/kg BW gentamicin, only the dose-dependent effects of iohexol were seen. Neither various plasma components nor light/electron microscopy showed any changes that could solely be related to the contrast medium. CONCLUSIONS: Iohexol produces transient renal effects in gentamicin nephropathy, which may be due to both chemotoxic and osmotic mechanisms.

Animals↗
PubMed1993

Low-molecular-mass proteinuria as a marker of proximal renal tubular dysfunction in normo- and microalbuminuric non-insulin-dependent diabetic subjects.

We determined the urinary excretion, expressed as the protein/creatinine ratio (morning urines), of albumin (a marker of glomerular dysfunction) and retinol-binding protein (RBP; a low-molecular-mass protein marker of tubular proteinuria) in 102 non-insulin-dependent diabetic patients. There was a statistically significant (P < 0.0001) correlation (rho = 0.38) between the urinary excretion values of the two proteins. The population could be divided into four subgroups: 32 with normal excretion values, 15 with above-normal urinary excretion of RBP, 24 with above-normal urinary excretion of albumin, and 31 patients with above-normal urinary excretion of both proteins. No patients had above-normal serum creatinine concentrations or above-normal serum RBP concentrations. This seems to exclude "tubular overflow proteinuria" as the cause of the increased urinary excretion of RBP seen in some patients with non-insulin-dependent diabetes. Our data suggest the presence of a state of proximal tubular dysfunction in these patients.

Adult↗
PubMed1992

Urinary excretion of albumin and retinol-binding protein in lithium-treated patients: a longitudinal study.

The urinary excretion of albumin and retinol-binding protein was determined by means of sensitive immunochemical methods in a 2-year longitudinal study of 22 lithium-treated patients. During lithium treatment, there were no significant changes in the median albumin:creatinine ratios or retinol-binding protein:creatinine ratios. However, the median albumin:creatinine ratios were significantly higher in lithium-treated patients than in 22 normal subjects, which indicates that glomerular permeability is increased; no correlation with serum lithium level or duration of lithium treatment could be shown. In patients treated with slow-release tablets, the urinary excretion of albumin was significantly higher after 2 years than in patients given lithium carbonate. The median retinol-binding protein:creatinine ratios did not differ significantly between lithium-treated patients and normal subjects, suggesting that lithium does not affect the catabolism of low-molecular-weight proteins in the proximal tubules. In conclusion, the study supports the hypothesis that lithium treatment produces a small but significant and nonprogressive elevation of the urinary excretion of albumin.

Adult↗
PubMed1991

Urine profiles and kidney histology following intravenous diatrizoate and iohexol in the degeneration phase of gentamicin nephropathy in rats. Effects on urine and serum profiles.

Urine chemical profiles were followed for three or nine days after intravenous injection of diatrizoate, iohexol, or saline in 30 rats, where a tubulointerstitial nephropathy was induced by gentamicin given over an eight-day period. Another ten rats injected with saline served as controls. Compared to injection of saline, both iohexol and diatrizoate induced dysfunction. The excretion of the cytoplasmic enzyme lactate dehydrogenase was significantly greater following iohexol than following diatrizoate. No significant differences between the two media were shown by the various serum components examined. Among the gentamicin-treated rats, light microscopy showed prolonged occurrence of tubular necrosis and a more intensive round cell infiltration following iohexol than following diatrizoate and saline. Both contrast media induced further temporary renal dysfunction in rats with gentamicin nephropathy; iohexol induced more morphologic changes than diatrizoate.

Animals↗
PubMed1991

Pretreatment with steroids before intravenous injection of diatrizoate or iohexol. Effects on urine and serum profiles.

The effects on urine and serum profiles of intravenous injection of diatrizoate, iohexol, or saline were studied in male rats pretreated with steroids or saline. Using urinary albumin, glucose, sodium, and the enzymes lactate dehydrogenase (LDH), gamma-glutamyltransferase (GGT), and N-acetyl-beta-D-glucosaminidase (NAG) as markers of glomerular and tubular function, it was found that diatrizoate caused temporary glomerular and tubular dysfunction; the effect was independent of the kind of pretreatment. Iohexol did not cause increased glomerular permeability in steroid- and saline-pretreated rats. When used following saline, iohexol induced increased excretion of three tubular components, whereas iohexol plus steroids caused increased excretion of all five tubular components. The dysfunctional effect of iohexol plus steroids was less than that of diatrizoate plus steroids. The serum components revealed no abnormalities induced by either contrast media or methylprednisolone. Pretreatment with steroids has no effect on the glomerular or tubular dysfunctional effect of diatrizoate, whereas it worsens the temporary tubular dysfunctional effect of iohexol in rats.

Acetylglucosaminidase↗
PubMed1991

Atrial natriuretic peptide and renal adaptation to contralateral nephrectomy in healthy man.

Atrial natriuretic peptide (ANP), angiotensin II (AII), aldosterone (Aldo) and arginine vasopressin (AVP) in plasma were determined in 12 healthy renal transplant donors before and 5, 12, 26, 54 days after uninephrectomy (Nx) in order to study the possible role of these hormones in functional adaptation to acute reduction in renal mass. Glomerular and tubular function was studied by measurements of the clearances of 51Cr-EDTA, lithium, sodium, potassium, and albumin. ANP was 7.4 +/- 3.1 pmol l-1 (mean +/- SD) before Nx and 8.7 +/- 6.1 pmol l-1 at 5 days after Nx and remained at this level through the observation period. Aldo showed a non-significant transient fall at 5 days after Nx. AII and AVP remained normal after Nx. At 5 days after Nx glomerular filtration rate (GFR) of the remaining kidney had risen from 45 +/- 7 ml min-1 before Nx to 57 +/- 8 ml min-1 (p less than 0.01), lithium clearance had risen from 13 +/- 2 ml min-1 before Nx to 20 +/- 7 ml min-1 (p less than 0.01), and sodium and water balance was normal. To conclude, plasma ANP, AII, Aldo and AVP do not appear to be responsible for the hyperfiltration and depression of fractional proximal sodium and water reabsorption observed in recently uninephrectomized man with normal sodium and water balance.

Absorption↗