Herbert Witzel

STRUCTURAL RELATIONSHIP BETWEEN THE MAMMALIAN FE(III)-FE(II) AND THE FE(III)-ZN(II) PLANT PURPLE ACID PHOSPHATASES

The primary structure of uteroferrin (Uf), a 35 kDa monomeric mammalian purple acid phosphatase (PAP) containing a Fe(III)‐Fe(II) center, has been compared with the sequence of the homodimeric 111 kDa Fe(III)‐Zn(II) kidney bean purple acid phosphatase (KBPAP). The alignment suggests that the amino acid residues ligating the dimetal center are identical in Uf and KBPAP, although the geometry of the coordination sphere might slightly differ. Secondary structure predictions indicate that Uf contains two βαβαβ motifs thus resembling the folding topology of the plant enzyme. Guided by the recently determined X‐ray structure of KBPAP a tentative model for the mammalian PAP can be constructed.

Crystallisation and preliminary analysis of glucose isomerase from Streptomyces albus.

The glucose isomerase of Streptomyces albus has been crystallised from a dilute solution of magnesium chloride buffered at a pH of 6.8–7.0. The crystals are in the space group I222 with cell dimensions a=93.9 Å, b=99.5 Å and c=102.9 Å. There is one monomer of the tetrameric molecule per asymmetric unit of the crystal and the packing density is 2.93 Å3·Da−1. The tetramer sits on the 222 symmetry point of the crystal. Native data have been recorded to a resolution of 1.9 Å and the crystals diffract to about 1.5 Å. The α‐carbon coordinates of the Arthrobacter glucose isomerase and the backbone coordinates of the S.olivochromogenes enzyme determined by other groups have been oriented in the present cell. The structure is currently being refined. The binding of several metal ions to the two metal sites has been analysed.

Steroid binding to the cytosolic estrogen receptor from rat uterus. influence of the orientation of substituents in the 17-position of the 8β- and 8α-series

The exact chemical and sterical requirements in the 17-position of 8β- and 8α-estrogens for an effective binding to the cytosolic receptor of immature rat uterus were investigated by competition experiments under non-equilibrium conditions. Oxygen or nitrogen functions with free electron pairs seem to be of essential importance in the 17-position. In contrast to 17α-methyl-, -vinyl- or -ethinyl-substituents a 17α-ethyl group strongly disturbs receptor binding. Also the introduction of a quasi equatorial allene or a 17β-ethinyl group reduces receptor binding. In comparison to the 8β -estrogens, the 8α-derivatives always showed lower, but still significant receptor binding and similar response to changes of substituents in the 17-position.

Crystallization and preliminary crystallographic data of purple acid phosphatase from red kidney bean

Purple acid phosphatase from red kidney bean has been crystallized from ammonium sulfate solutions in the pH range from 3.5 to 5.5. The crystal form is tetragonal bipyramidal and the largest crystals grew up to 2.0 mm long. Systematic absences indicate one of the enantiomorphic space groups P4(1)2(1)2 (92) or P4(3)2(1)2 (96) with cell dimensions a = b = 104.1(1) A and c = 308.7(2) A. The asymmetric unit contains one dimer with Mr of 110,700, determined by ultraviolet-laser desorption mass spectrometry. The crystals, with a salt-free density of 1.12 g/cm3 and a water content of 67%, diffract to 3.5 A.

Coenzyme A Glutathione Disulfide A Potent Vasoconstrictor Derived From the Adrenal Gland

The adrenal gland is involved in the regulation of vascular tone by secretion of vasoactive agents such as catecholamines, neuropeptide Y, or endogenous ouabain. A further potent vasoconstrictor is isolated from bovine adrenal glands and is identified by chromatography, mass spectrometry, UV spectroscopy, and enzymatic cleavage as coenzyme A glutathione disulfide (CoASSG). CoASSG is found in chromaffin granules of adrenal glands and is released from adrenal medulla slices by carbachol. At a concentration of 10(-12) mol/L CoASSG increases renal vascular resistance. Intra-aortic injection of 5 x 10(-10) mol CoASSG increases blood pressure in the intact animal. Besides its vasopressor properties, this substance potentiates the effects of angiotensin II on vascular tone. It is concluded that CoASSG could play a role in blood pressure regulation not only by direct effects but also by modulation of the action of angiotensin II.

Effect of plasma from essential hypertensives on vascular tone of aortic strips, isolated perfused mesentery and isolated perfused kidney

Different vascular models of normotensive Wistar rats, including aortic strips, isolated perfused mesentery and isolated perfused kidney, were used to study hemodynamic effects of plasma fractions obtained by gel filtration from the blood of essential hypertensive and normotensive subjects. Plasma fractions from essential hypertensives studied had been shown to increase blood pressure after intravenous injection in rats. In the aortic strips, 50 microliters of a hypertensive fraction (HF) elicited a calcium-dependent contraction of 0.14 +/- 0.035 mN (n = 20, p less than 0.05), which was inhibited by nifedipine, whereas tension of the strips was not significantly changed by normotensive fractions (NF) (n = 17). In the isolated perfused mesentery preparation, no significant change of perfusion pressure by HF or NF could be demonstrated (n = 10). In the isolated perfused kidney, a transient increase of perfusion pressure was induced by HF (19.5 +/- 16.6 mm Hg, n = 40, P less than 0.001) but not by NF. This increase was abolished in calcium-free, 2 mmol/l EGTA containing perfusion medium. The response was diminished, but not abolished by nifedipine. These data demonstrate vasopressor properties of plasma from essential hypertensives, which might be the consequence of a circulating vasoconstrictor substance in the blood of essential hypertensives.

Shielding effects at 17α-substituted estrogens. A tentative explanation for the low biological activity of 17α-ethyl-estradiol based on I.R. and NMR spectroscopic studies

Abstract Infrared data of the hydroxyl stretching band and NMR data of the hydroxyl proton for 7 different 17-substituted estradiol-3-methyl-ether compounds have been recorded. The band positions can be related to the extent of shielding effects or intramolecular interactions of hydrogen bonding type. The splitting of several i.r. bands can be explained on the basis of rotamers and restrictions in the free rotation of the hydroxyl group. This holds especially for 17α-ethyl-estradiol, in which the access to the free electron pairs of the OH group is hindered by the 17α-ethyl group. This may explain the very low receptor binding and reduced biological activity of 17α-ethyl-estradiol in contrast to the stronger binding of 17α-methyl-17α-vinyl or 17α-ethinyl-estradiol.

Effect of Plasma from Essential Hypertensives on Tension of Aortic Strips

The effect of fractions of plasma from essential hypertensive (n = 27) and normotensive subjects (n = 26) on the tension of aortic strips (n = 27) from normotensive rats was examined. The fractions obtained from hypertensive patients had been shown to increase blood pressure, when injected intravenously in a normotensive rat. In aortic strips the hypertensive fractions elicited a transient relaxation of variable amplitude and subsequently a sustained contraction. The normotensive fractions did not alter tension of the strips significantly. In Ca2+ free medium and after addition of nifedipine hypertensive plasma fractions did not induce a contraction, whereas in Na+ free medium the contraction was not abolished. It is concluded that in the fraction of hypertensive plasma containing substances with a molecular weight in the range of 1000-1500 Da a vasopressor agent with direct actions on arterial smooth muscle is present. The substance probably acts by increasing Ca2+ influx in vascular smooth muscle.

Cobalt X-ray absorption spectroscopy of glucose isomerase

Introduction Glucose isomerases catalyze the isomerization of D-glucose to D-fructose and of D-xylose to D-xylulose. They are of considerable interest in the industrial production of high-fructose corn syrup. Glucose isomerase from Streptomyces albus (GI) is a tetramer (Mr = 172.000) with four identical subunits [1,2]. Each subunit contains two distinct metal bindin of Co2+, Mn f sites, a tight binding site (B-site) and a weaker binding site (A-site). Addition + or Mg2’ * increases the catalytic activity as well as the stability of the enzyme.

Stable nitrones from 17β-hydroxylamine derivatives of 8β- and 8α-estrone

17β-Hydroxylamine derivatives of 8β and 8α-estrone can be obtained by reduction of the corresponding oximes with diborane or NaCNBH3. The products can be converted easily and reversibly to nitrones by addition of aldehydes or ketones. The nitrones are more stable than the corresponding hydroxylamines, and may be used as protecting groups.