l-Phenylalanine active transport in the midgut of Bombyx mori larva

l-phenylalanine and α-aminoisobutyric acid are actively transported from the lumen to the haemolymph in the isolated midgut of Bombyx mori larva. Kinetics of l-phenylalanine fluxes as a function of concentration have been studied. The influx shows a convex relationship to the aminoacid concentration, while the outflux bears a linear relationship within the range of concentrations considered. The net flux shows a saturation kinetic, is sodium independent and is inhibited by DNP and anoxia. l-phenylalanine pools obtained by luminal or haemolymph loading are similar and no intracellular accumulation of the aminoacid takes place. Conversely the amount of α-aminoisobutyric acid entering the cells through the basolateral membrane exceeds that of luminal origin and it reaches an intracellular concentration twofold higher than that of the bathing medium.     

Density-labelling studies on the photocontrol of phenylalanine ammonia lyase levels in Cucumis sativus

The de novo synthesis of PAL is demonstrated to occur sometime between imbibition and the end of a 4-hr white light treatment. H₂OD₂O transfer experiments indicate that PAL synthesis may occur during the light period whilst D₂O-H₂O transfer experiments indicate that synthesis of inactive PAL may occur during dark growth followed by activation by light. Neither of these observations is conclusive. De novo synthesis of PAL occurs in excised hypocotyls of gherkin and tuber discs of potato either in darkness or in light. It is concluded that there is as yet no evidence which definitively shows that light controls PAL levels by regulating the rate of de novo synthesis.     

l-Proline transport in Saccharomyces cerevisiae

Transport of l-proline into Saccharomyces cerevisiae K is mediated by two systems, one with a K_T of 31 μM and J_max of 40 nmol · s^?1 · (g dry wt.)^?1, the other with K_T > 2.5 mM and J_max of 150–165 nmol · s^?1 · (g dry wt.)^?1, The kinetic properties of the high-affinity system were studied in detail. It proved to be highly specific, the only potent competitive inhibitors being (i) l-proline and its analogs l-azetidine-2-carboxylic acid, sarcosine, d-proline and 3,4-dehydro-dl-proline, and (ii) l-alanine. The other amino acids tested behaved as noncompetitive inhibitors. The high-affinity system is active, has a sharp pH optimum at 5.8–5.9 and, in an Arrhenius plot, exhibits two inflection points at 15°C and 20–21°C. It is trans-inhibited by most amino acids (but probably only the natural substrates act in a trans-noncompetitive manner) and its activity depends to a considerable extent on growth conditions. In cells grown in a rich medium with yeast extract maximum activity is attained during the stationary phase, on a poor medium it is maximal during the early exponential phase. Some 50–60% of accumulated l-proline can leave cells in 90 min (and more if washing is done repeatedly), the efflux being insensitive to 0.5 mM 2,4-dinitrophenol and uranyl ions, to pH between 3 and 7.3, as well as to the presence of 10–100 mM unlabeled l-proline in the outside medium. Its rate and extent are increased by 1% d-glucose and by 10 μg nystatin per ml.     

Growth of Rhodopseudomonas capsulata on l- and d-malic acid

d-malate replaced l-malate in supporting both photosynthetic (anaerobic, light) and heterotrophic (aerobic, dark) growth of Rhodopseudomonas capsulata. Growth rates and cell yields were nearly equivalent with both enantiomorphs. Addition of glucose to malate culture media increased the growth rate and doubled the cell yield of heterotrophic cultures, but had little effect on photosynthetic cultures. Aerobically-grown cells showed a higher level of substrate-dependent oxygen uptake with l-malate than with d-malate. This preference for l-malate occured even in cells grown on d-malate. No malic racemase activity was detected in extracts of heterotrophically- or photosynthetically-grown cells.     

Synthesis of aminocyclitols from l-quinic acid

A variety of 1,3-diamino and 1,4-diaminocyclitols, manoaminocyclitols, and triaminocyclohexanol have been synthesized starting with the chiral ketone intermediate, 2, derived from l-quinic acid. Reduction of 2 with lithium borohydride afforded two epimeric diols (4 and 5), both of which were transformed by straight-forward but distinctly different chemical procedures into potentially useful aglycons for preparing novel tupes of bioactive, aminocyclitol glycoside antibiotics. The disposition of the substituents at C-1, C-3, C-4, and C-5 in 19 and 37 is identical with that present in the 2-deoxystreptamine nucleus in the naturally occurring antibiotics     

Novel biosynthesis of d-pinitol in simmondsia chinensis

Chase experiments with ¹⁴CO₂ and feeding experiments with labelled inositols showed that d-pinitol in leaves of Simmondsia chinensis arises via epimerization of d-ononitol. This finding represents an alternative pathway, since d-pinitol is formed in gymnosperms and other plants by epimerization of sequoyitol.     

Alternate pathways of l-rhamnose transport in Arthrobacter pyridinolis

Phosphoenolpyruvate: hexose phosphotransferase-negative mutants of Arthrobacterpyridinolis fail to grow on l-rhamnose. Although phosphoenolpyruvate: l-rhamnose phosphotransferase activity could not be consistently demonstrated in extracts of rhamnose-grown cells, low levels of phosphoenolpyruvate-dependent uptake of rhamnose were found using isolated membrane vesicles from rhamnose-grown cells. This uptake was not inhibited by uncoupling agents or an inhibitor of the respiratory chain. Phosphotransferase-negative mutants could grow on l-rhamnose if l-malate was also present in the medium. l-Malate- and succinate-dependent uptake of rhamnose was found in membrane vesicles. Either of the two oxidizable substrates caused a 5-fold stimulation of the rate of l-rhamnose uptake over that observed in the absence of additions. Malate-dependent l-rhamnose uptake had a K_m for rhamnose of 2.9 × 10^?6m. It was inhibited by uncoupling agents, inhibitors of the respiratory chain, and sulfhydryl reagents.     

Active transport of l-sorbose and 2-deoxy-d-galactose in Saccharomyces fragilis

Sorbose and 2-deoxy-d-galactose are taken up in Saccharomyces fragilis by an active transport mechanism, as indicated by the energy requirement of the process and the accumulation of free sugar against the concentration gradient. There are no indications for transport-associated phosphorylation as mechanism of energy coupling with these two sugars.The measured sugar-proton cotransport and the influx inhibition by uncouplers suggest a chemiosmotic coupling mechanism. Thus there are at least two different active transport mechanisms operative in Saccharomyces fragilis: transport-associated phosphorylation in the case of 2-deoxy-d-galactose and chemiosmotic coupling in the case of sorbose and 2-deoxy-d-galactose. The difference between the two mechanisms are discussed.Uncouplers do not stimulate downhill sorbose transport in energy-depleted cells and evoke an almost complete inhibition of efflux and of exchange transport.The differences between this sugar-proton cotransport system and similar systems in bacteria and Chlorella are discussed.     

Solubilization and properties of the enzyme-cleaving 13-l-hydroperoxylinolenic acid in tea leaves

The membrane bound hydroperoxide lyase (E″₂) which catalyses the cleavage of 13-l-hydroperoxides (18:3-OOH and1 8:2-OOH) of linolenic and lin     

CNDO/2 calculations of the relative stability of poly (l-proline I) and poly (l-proline II)

The CNDO/2 method using the tight binding approximation for polymers was applied to poly(l-proline I) and poly(l-proline II). The calculations were also carried out for poly(l-alanines) and model molecules which have the same backbone geometrics as those of poly(l-prolines). The results obtained show that both forms of poly(l-proline I) and poly(l-proline II) have nearly the same energy in agreement with experimental results. From the analysis of the total energy, it was found that the intrasegment energy of poly(l-proline II) was lower than that of poly(l-proline I) while the intersegment energy of poly(l-proline I) was lower than that of poly(l-proline II). This result can be considered to correspond well with the experimental fact that poly(l-proline II) is more stable in good or polar solvents and poly(l-proline I) in poor or non-polar solvents. The analysis of the total energy of poly(l-proline) leads us to the conclusion that the α and β carbons play an important role in determining the relative stability between poly(l-proline I) and poly(l-proline II) and the γ carbon does hvae a marked effect on the electronic structures of the polymers in question. This conclusion was also confirmed by comparison of the electronic structures of poly(l-prolines) with those of poly(l-alanines) and model compounds concerned.     

Effect of nitrogen sources upon the activity of l-glutamate dehydrogenase of Lemna gibba

When Lemna gibba cultures, grown on medium containing l-glutamate as the sole nitrogen source are transferred to medium in which ammonium is the only source of nitrogen, the activity of a NAD-dependent l-glutamate dehydrogenase (GDH) increases approximately 5-fold over 3 days. Upon re-transfer to glutamate medium the activity declines to its initial value after a further 6 days. The rise in activity is independent of the presence of EDTA and is not the result of an increase in the ease with which the enzyme can be extracted. p-Fluoro-dl-phenylalanine, azetidine-2-carboxylic acid and puromycin but not d-threo-chloramphenicol, erythromycin or lincomycin inhibit the increase when included in ammonium medium. These observations, together with those obtained from the use of a deuterium oxide-labelling technique, suggest that the increase in GDH activity is due to de novo synthesis on 80S ribosomes.     

Formation of glycine and aminoacetone from l-threonine by rat liver mitochondria

Threonine is a precursor of glycine in the rat, but the metabolic pathway involved is unclear. To elucidate this pathway, the biosynthesis of glycine, and of aminoacetone, from l-threonine were studied in rat liver mitochondrial preparations of differing integrities. In the absence of added cofactors, intact mitochondria formed glycine and aminoacetone in approximately equal amounts from 20 mM l-threonine, but exogenous NAD⁺ decreased and CoA increased the ratio of glycine to aminoacetone formed. In intact and freeze-thawed mitochondria, the ratio of glycine to aminoacetone formed was markedly sensitive to the concentration of l-threonine, glycine being the major product at low l-threonine concentrations. Disruption of mitochondrial integrity by sonication (1 min) decreased the ratio of glycine to aminoacetone formed, and in 20 000 × g supernatant fractions from sonicated (3 min) mitochondria, aminoacetone was the major product. The main non-nitogenous tow-carbon compound detected when intact mitochondria catabolized l-threonine to glycine was acetate, which was probably derived from deacylation of acetyl-CoA. These results suggest that glycine formation from l-threonine in rat liver mitochondria occured primarily by the coupled activities of threonine dehydrogenase and 2-amino-3-oxobutyrate CoA-ligase, the extent of coupling between the enzymes being dependent upon a close physical relationship and upon the flux through the dehydrogenase reaction. In vivo glycine synthesis would predominate, and aminoacetone would be a minor product.     

Detection and properties of l-histidinol dehydrogenase in wheat germ

The presence and partial characterization of the properties of l-histidinol dehydrogenase (EC 1.1.1.23), the enzyme catalysing the last step in the pathway of histidine biosynthesis, has been described in higher plants for the first time. The activity has been found in cell-free extracts from wheat germ, turnip root, radish root and squash fruit. The enzyme has been partially purified and characterized from extracts of acetone powders of wheat germ. DEAE-cellulose chromatography revealed two peaks of histidinol dehydrogenase activity. In one there was a rapid reduction of NAD⁺ in the absence of histidinol; however, the rate was stimulated by the addition of histidinol. The rate in the absence of substrate became quite low after several min and the histidinol-dependent rate was then easily observed. The second peak of activity did not reduce NAD⁺ unless l-histidinol was present in the assay mixture. The K^ms for l-histidinol and NAD⁺ were determined for this latter enzyme. The values obtained at saturating concentrations of the other substrate were l-histidinol, 8.8 μM and NAD⁺, 0.14 mM. The product of the dehydrogenase reaction was histidine as determined by paper chromatography.     

l-2-Oxalylamino-3-aminopropionic acid,an isomer of Lathyrus sativus neurotoxin

The first chemical synthesis of l-2-oxalylamino-3-aminopropionic acid, an isomer of the Lathyrus sativus neurotoxin, is described. Studies on its biological properties are reported. Experiments with l-3-[¹⁴C-oxalyl]amino-2- aminopropionic acid show that the amount of 2-oxalylamino isomer detectable in seed extracts can be accounted for by rearrangement which occurs during isolation.     

An S-adenosyl-l-methionine; coniine methyltransferase from Conium maculatum

Evidence is presented for a cell free system from Conium maculatum which catalyses the transfer of a methyl group from S-adenoysl-l-methionine to coniine with the formation of N-methyl coniine. Maximum enzyme activity which occurred in the unripe fruits was enhanced by dithiothreitol, and evidence for the role of sulphydryl groups of the enzyme was obtained from inhibition with p-CMB, iodoacetamide and N-methyl maleimide. A divalent metal cation dependency was not detected.     

Simultaneous determination of d- and l-thyroxine in human serum by liquid chromatography with electrochemical detection

A method for the determination of d- and l-thyroxine in human serum is described. The method involves extraction of thyroxine from serum and the separation of thyroxine enantiomers on a reversed-phase, high-performance liquid chromatographic column by use of a chiral eluent containing l-proline and cupric sulfate. Satisfactory resolution of the enantiomers of thyroxine, triiodothyronine, and reverse triiodothyronine can be achieved in 12 min and, employing amperometric detection to monitor the separation, the detection limit for serum thyroxine is in the range of 1–3 ng per injected sample.     

Mode of action of a xylanase and its significance for the structural investigation of the branched l-arabino-d-glucurono-d-xylan from redwood (Sequoia sempervirens)

The substitution pattern of the water-soluble l-arabino-(4-O-methyl-d-glucurono)-d-xylan from redwood (Sequoia sempervirens) has been studied by enzymic degradation. Exhaustive hydrolysis by an endo-xylanase (EC 3.2.1.8) from a Basidiomycete Sporotrichum dimorphosporum left a residue accounting for 20% of the original d-xylan. In the dialyzable material, oligosaccharides having arabinose or 4-O-methylglucuronic acid residues attached to the non-reducing d-xylosyl end-group of xylobiose or xylotriose, respectively, were the smallest branched oligomers released. Action of the xylanase appears to involve a region of the polysaccharide backbone having three xylosyl residues. A mode of action is proposed that requires unsubstituted hydroxyl groups at C-2, C-3, and C-2′ of a xylobiosyl residue. The binding site seems to correspond to a shallow cavity. The composition and structure of the final residue of attack shows that the enzyme has no action when the xylosyl residues branched through O-2 are separated by only one, unsubstituted xylose residue. This pattern of action, the nature of the dialyzable products, and the production of a final residue in which the substituents are accumulated, suggest that the arabinosyl and glucosyl-uronic groups are irregularly distributed on the main chain of the xylan from redwood and that in some regions they are in close vicinity when not actually on adjacent xylosyl residues.     

Kinetic study of the pathway of melanizationn between l-dopa and dopachrome

The first part of the melanization pathway from l-dopa to dopachrome has been studied as a system of various chemical reactions coupled by an enzymatic reaction. A theoretical and experimental kinetic approach is proposed for such a system. Rate constants for the implicated chemical steps at different pH and temperature values can be evaluated from measurement of the lag period arising from the accumulation of dopachrome that takes place when l-Dopa was oxidized at acid pH. The thermodynamic parameters of the chemical steps, the deprotonation of dopaquinone-H⁺ into dopaquinone and the internal cyclization of dopaquinone into leukodopachrome, have been obtained. From the results presented, an alternative series of chemical reactions to the Raper-Mason scheme are proposed and discussed.     

Differences in the incorporation pattern of labelled cinnamic acid and l-phenylalanine into the anthocyanins of Petunia hybrida

The incorporation of phenylalanine-[¹⁴C] into anthocyanins of petals of Petunia hybrida is greater than that of cinnamic acid-[¹⁴C]. Moreover, there is a preferential incorporation of phenylalanine-[¹⁴C] into delphinidin 3-monoglucoside, as compared with the incorporation into cyanidin and peonidin 3-monoglucosides.     

Azobenzene-containing poly(l-glutamates). Photochromism and conformation in solution

Photochromic polypeptides having various contents of azobenzene chromophores attached to th side chains have been prepared by condensing poly(l-glutamic acid) with p-amino-azobenzene. The photochemical and thermal cis-trans isomerizations of the azo chromophores have been investigated by absorption and circular dichroism spectroscopy, and the photochromic behaviour has been related to the conformation in solution. The azopolypeptides exhibit the α-helix c.d. pattern, in trimethylphosphate. The α-helix content markedly depends on the azo content, but it is not affected by the cis or trans geometric forms of the azo side chains. Strong solvent effects by H₂O or trifluoroethanol on the extrinsic azo c.d. bands, suggest the existence, in trimethylphosphate solution, of a super-ordered secondary structure involving a regular arrangement of the side chains, on th periphery of the helical peptide backbone.