Inhibition of Aspartate Aminotransferase by Glycation In Vitro Under Various Conditions

Incubation of 50 mM d -glucose with aspartate aminotransferase (AST, EC 2.6.1.1) preparations (purified pig heart enzyme or a rat liver 20,000 × g supernatant) at 25°C had no effect on enzyme activity. 50 mM d -fructose or d -ribose gradually inhibited pig heart AST under the same conditions to zero activity after 14 days. 50 mM dl -glyceraldehyde decreased enzyme activity to zero after 6 days of incubation. The inhibition of pig heart AST by 50 mM d -fructose or d -ribose was marked even at a temperature of 4°C but it was less pronounced than at 25°C. There was no effect of 0.5 mM 2-oxoglutarate on AST activity during incubation, while the presence of 25 mM l -aspartate decreased it rapidly. 0.5 mM 2-oxoglutarate partly prevented inhibition of AST by d -ribose or d -fructose, while an analogous experiment with 25 mM aspartate resulted in a rapid decline similar to that in the absence of sugars.     

A severe autosomal recessive acromesomelic dysplasia,the Hunter-Thompson type,and comparison with the Grebe type

Summary Two siblings with a short-limb dwarfing condition which we call acromesomelic dysplasia, Hunter-Thompson type are reported. Abnormalities are limited to the limbs and limb joints in this severe form of dwarfism. The middle and distal segments of the limbs are most affected. The lower limbs are more affected than the upper. We are aware of one previously published case of this entity reported by A. G. W. Hunter and M. W. Thompson in 1976. Dislocations of the elbows and ankles were present in all three patients and dislocations of the hips and knees in two. One of the siblings who did not have hip and knee dislocations clinically resembled Grebe chondrodysplasia, another severe acromesomelic dwarfing condition. However, radiological analysis suggests that while acromesomelic dysplasia, Hunter-Thompson type and Grebe chondrodysplasia are related, they are not identical. Grebe chondrodysplasia has been established as an autosomal recessive trait. It appears probable that the entity we describe has the same mode of genetic transmission.     

Thermal melting and circular dichroism of DNA complexes with (Lys-Ala-Ala)10 and (Lys-Ala-Ala)n: effect of polypeptide chain length

DNA complexes with polypeptides (Lys-Ala-Ala)₁)] and (Lys-Ala-Ala)₃₄ have been studied using the methods of thermal melting and circular dichroism. Derivative melting curves of (Lys-Ala-Ala)₁₀ DNA differed substantially from those of (Lys-Ala-Ala)₃₄ prepared either by salt gradient dialysis or by direct mixing. Melting curves of the former complex were unimodal or bimodal with T_m increasing continuously withn input lysin-to-DNA phosphate ratio (r); those of the latter complex consisted of three separate transitions with T_m values almost independent of r. Complete reversibility of binding in the (Lys-Ala-Ala)₁₀-DNA system but a slow redistribution of (Lys-Ala-Ala)₃₄ on DNA at low temperature were found in the redistribution experiments Much faster redistribution from denatured to native DNA occurs at the temperature of melting, contributing to the unusual trimodal melting pattern. Circular dichroism curves are very similar for both complexes and indicate little change of DNA conformation upon polypeptide binding.     

Studien über die Funktion der im Wasser gelösten Nährsubstanzen im Stoffwechsel der Wassertiere

Ohne Zusammenfassung     

External factors involved in the regulation of an extracellular proteinase synthesis in Bacillus megaterium

Summary A mathematical model was formulated to describe the kinetics and stoichiometry of growth and proteinase production in Bacillus megaterium. Synthesis of the extracellular proteinase in a batch culture is repressed by amino acids. The specific rate of formation of the enzyme (r _E) can be described by the formula {ie373-1}, where k ₂ and k ₃ stand for the non-repressible and repressible part of enzyme synthesis respectively, k _{S 2} is a repression coefficient and S ₂ indicates the concentration of amono acids; the values of k ₂ and k _{S 2} depend on the composition of the mixture of amino acids. Even in a high concentration, a single amino acid is less effective than a mixture of amino acids. The dependence of the proteinase repression on the concentration of an external amino acid (leucine) follows the same course as its rate of incorporation into proteins, approaching saturation at concentrations higher than 50 M (half saturation approximately 10 M). However, the total uptake of leucine did not exhibit any saturation even at 500 M external concentration.Symbols X biomass concentration, g/l - E proteinase concentration, unit/l - t time, h - S ₁ concentration of glucose, g/l - S ₂ concentration of amino acids, g/l - specific growth rate, l/h - rE specific rate of enzyme production, unit/g/h - k ₁ growth kinetic constant, l/h - k ₂ product formation kinetic constant (for non-repressible part of enzyme synthesis), unit/g - k ₃ product formation kinetic constant (for repressible portion of enzyme synthesis), unit/g - k _{S 1} saturation constant, g/l - k _{S 2} repression coefficient for a certain amino acid or amino acids mixture, g/l     

Regulation of branched-chain amino acid biosynthesis in Streptomyces fradiae,a producer of tylosin

Synthesis of threonine dehydratase in Streptomyces fradiae was positively influenced by valine and negatively by isoleucine. However, these two amino acids had no effect on the activity of this enzyme. Synthesis of threonine dehydratase in -aminobutyrate resistant mutants of S. fradiae was pronouncedly less sensitive to the positive effect of valine and this change in regulation led to valine overproduction. Synthesis of acetohydroxy acid synthase is regulated in a similar manner to that of threonine dehydratase, however a lower level of expression was detected in -aminobutyrate resistant mutants. And again, no effect of branched-chain amino acids on acetohydroxy acid synthase activity was observed. It follows that in S. fradiae synthesis of threonine dehydratase is the main regulatory mechanism governing production and the mutual ratio of synthesized valine and isoleucine.Abbreviations -AB -aminobutyrate - AHAS acetohydroxy acid synthase - -KB -ketobutyrate - MNNG N-methyl-N-nitro-N-nitrosoguanidine - TD threonine dehydratase - Trans. B. transaminase of branched-chain amino acids - VDH valine dehydrogenase     

Carnitine transport and exogenous palmitate oxidation in chronically volume-overloaded rat hearts

L-Carnitine transport and free fatty acid oxidation have been studied in hearts of rats with 3-month-old aorto-caval fistula. For carnitine transport experiments, the hearts were perfused via the ascending aorta with a bicarbonate buffer containing 11 mM glucose and variable concentrations L-[14C]carnitine (10-200 microM). In some experiments, the active component of carnitine transport was suppressed by the adjunction of 0.05 mM mersalyl acid. The subtraction of passive from total transport allowed reconstruction of the saturation curves of the carrier-mediated transport of L-carnitine. Our data suggest that at a physiological carnitine concentration (50 microM), the rate of [14C]carnitine accumulation was significantly depressed in mechanically overloaded hearts. In addition, according to Lineweaver-Burk analysis, the affinity of the membrane carrier for L-carnitine was considerably diminished (Km carnitine 125 instead of 83 microM, Vmax unchanged). The above alterations of L-carnitine transport did not result from a decrease of the transmembrane gradient of sodium, since the intracellular Na+ content of the hypertrophied hearts was quite similar to that of control hearts. The ability of atrially perfused, working hearts to oxidize the exogenous free fatty acids was assessed from 14CO2 production obtained in the presence of [U-14C]palmitate or [1-14C]octanoate. The total 14CO2 production, expressed per min per g dry weight, was significantly diminished in hearts from rats with the aorto-caval fistula if 1.2 mM palmitate was used. On the other hand, in the presence of 2.4 mM octanoate, a substrate which circumvents the carnitine-acylcarnitine translocase, no such reduction of the 14CO2 production could be detected. Our results suggest that the decrease of L-carnitine transport, resulting in a significant depression of tissue carnitine, may impair long-chain fatty acid activation and/or translocation into mitochondria. In contrast, the oxidation of short-chain fatty acids, the activation of which takes place directly in mitochondrial matrix, is not limited in volume-overloaded hearts.     

Functional reconstitution of rat ovarian LH/hCG receptor into proteoliposomes

Rat ovarian membrane LH/hCG receptor was solubilized in various detergents and reconstituted into proteoliposomes. Upon removal of sodium cholate by active absorption on Bio-Beads SM-2, the functional interaction between receptor and adenylate cyclase was restored. Adenylate cyclase was stimulated by hCG, HCG+GTP or GppNHp and NaF. Reconstituted proteoliposomes bound more ¹²⁵I-hCG (528 fmol/mg protein) than membrane-bound receptors (384 fmol/mg protein). There was no difference, however, in the relative affinity of reconstituted receptor preparations for hCG.     

Catecholaminergic and peptidergic nerve components of intramural ganglia in the rat heart

Summary Immunohistochemical properties of the terminal nerve network in the rat heart were assessed by use of the elution-restaining method. The colocalization of the enzymes involved in catecholamine synthesis (tyrosine hydroxylase — TH, dopamine--hydroxylase — DBH) as well as the respective distributions of the neuropeptides associated with the adrenergic nervous system (neuropeptide tyrosine — NPY, C-terminal flanking peptide of neuropeptide Y — C-PON) were studied in series of serial sections throughout the interatrial septum and the atrioventricular junction. Our data suggest that ganglion cells of sulcus terminalis as well as the epicardial ganglia enclosed between the superior vena cava and ascending aorta are VIP- and TH-negative, but neuropeptide Y- and DBH-immunoreactive. They give rise to three intraseptal nerves directed towards the specialised structures of the atrioventricular junction. These nerve fascicles contain abundant, thick TH-immunoreactive nerve fibres and scarce, thin NPY- and DBH-immunoreactive fibres. The cell bodies of the intramural ganglion cells localized between the right and left branches of the bundle of His (Moravec and Moravec 1984) are strongly TH- and DBH-immunoreactive. They are innervated by thick nerve fibres having the same immunohistochemical properties (NPY- and DBH-immunoreactivities) as those of a subpopulation of the epicardial ganglion cells and seem to supply some of the TH-immunoreactive nerve fibres directed via the intraseptal nerves to the epicardial ganglia. The existence of a multicomponent nerve network, characterized by a reciprocal innervation of the sinus node and atrioventricular node areas, is suggested by our immunohistochemical data.