乳酸菌酸胁迫反应机制研究进展

乳酸菌可发酵糖类产生乳酸,并广泛应用于食品、药物和饲料等工业。由于有机酸的积累,乳酸菌大部分的生长代谢都在低pH的酸性环境中进行,具有酸胁迫反应。pH的自我平衡、ATR反应机制、对大分子的保护和修复作用及细胞膜的变化等是乳酸菌酸胁迫反应的主要机制,其中,pH自我平衡包括F0F1-ATPase质子泵、精氨酸脱氨酶途径(ADI)和谷氨酸脱羧酶途径(GAD)等。由此可见,乳酸菌酸胁迫反应机制涉及到基因和蛋白的表达调控等,是非常复杂的网络调控体系。     

Purification and Characterization of the Heat-Stable Factors Essential for the Conversion of Lignoceric Acid to Cerebronic Acid and Glutamic Acid: Identification of N-Acetyl-l-Aspartic Acid

Abstract: The conversion of lignoceric acid to cerebronic acid, ceramides, cerebrosides, and glutamic acid is catalyzed by a rat brain particulate preparation. The heat-stable factor, prepared from calf cerebellum, together with the heat-labile factor, a pyridine nucleotide, and Mg²⁺ are essential to all of these metabolic pathways. Our previous work showed that the heat-stable factor is composed of at least two components, HSF-1 and HSF-2, and identified HSF-2 as d -glucose-6-phosphate. In the current investigation, HSF-1 was further purified and found to be N -acetyl- l -aspartic acid. In addition, it was discovered that a third component, HSF-3, is also required for heat-stable factor activity. A reconstituted system composed of N -acetylaspartic acid, glucose-6-phosphate, and HSF-3 fully replaced the heat-stable factor essential for the conversion of lignoceric acid to cerebronic acid and glutamic acid. The reconstituted heat-stable factor did not show the initial time lag always observed with the crude heat-stable factor.     

Oxidation of Nicotinic Acid by a Bacillus Species: Regulation of Nicotinic Acid and 6-Hydroxynicotinic Acid Hydroxylases

The first two enzymes employed by a Bacillus species for the dissimilation of nicotinic acid are coordinately induced. The inducer of the enzymes appears to be 6-hydroxynicotinic acid, the product of the first enzyme in the pathways. Synthesis of the enzymes is repressed by glucose when ammonium is present in the medium, but not when nicotinic acid is the sole nitrogen source. The possible significance of the coordinate induction and unusual repression is discussed.     

Ultraviolet-Induced Cross-Links in the Deoxyribonucleic Acid of Single-Stranded Deoxyribonucleic Acid Viruses as a Probe of Deoxyribonucleic Acid Packaging

Deoxyribonucleic acid (DNA) from ultraviolet (UV)-irradiated phiX174 sediments in alkali at rates up to 1.7 times that of unirradiated phiX174 DNA and is observed as a condensed, cross-linked structure when examined in the electron microscope by the formamide spreading technique. This structure appears to result from multiple cross-links induced in the tightly coiled DNA contained within the spherical phiX174 capsid. In contrast, the DNA extracted after UV irradiation of the filamentous bacteriophage M13 is not strikingly altered in its sedimentation properties and appears by electron microscopy to be rod-shaped as a result of side-to-side association of the circular DNA. The differences in these UV-induced structures reflect the differences in the packaging of the single-stranded DNA in the two virions.     

The Hazard of Acid Differentiation in Gomori's Method for Acid Phosphatase

In his original method for the histochemical demonstration of acid phosphatase, Gomori prescribed differentiation of incubated sections by rinsing them in 14% aqueous acetic acid, to remove the nonspecifically precipitated lead deposits. According to him, the enzymatically produced lead phosphate is not washed out by this procedure. As a result of recent improvements in tissue preparation and shorter incubation time, this staining reaction as it is used now is quite sensitive to an acetic acid wash. If this wash is used as recommended originally, it may completely abolish a truly positive reaction. To avoid falsely negative results, and to compare sections of normal and pathological tissue, omission of this differentiation by acetic acid is essential. The risk of mistaking nonspecific lead precipitates in the interpretation of a positive reaction is very small, and can be avoided by running a negative control slide in which no lead phosphate can be produced enzymatically.     

Hyaluronic Acid Degradation by Ascorbic Acid and Influence of Iron

The effects of ascorbic acid, iron and ADP on hyaluronic acid, a compound present in inflamed joints, were investigated in an in vitro system. Ascorbic acid induces degradation of hyaluronic acid which increased in the presence of FeCl, and which is additionally stimulated by ADP chelated ferric ions. The hyaluronic acid degrading reactions induced by the Fe-III/ADP/ascorbic acid system were inhibited by catalase and formate to various extents whereas the presence of superoxide dismutase did not exert any inhibitory effect. Desferrioxamine, a specific iron chelator, completely inhibited hyaluronic acid depolymerisation by ascorbic acid as well as in combination with FeCl₃ or FeCl₃/ADP, respectively. We suggest that the ultimate hyaluronic acid degrading species is OH', generated via the Fe-III/ADP catalysed Haber Weiss reaction. There is also an indication for the involvement of perferryl or/and ferryl species in the degradation process.     

Synthesis of Polyoxamic Acid (2-Amino-2-deoxy-l-xylonic Acid

The rate of precipitation of the retrograded amylose product from a dil. amylose solution was determined by the centrifugal method. The results showed that the relation of the quantity of precipitate vs. time did not fit the typical second order reaction for the coalescence of colloidal particles but fitted the crystallization formula, in appearance.

The rate of precipitation was in proportion to (c-c_a)^1.5, where c is the amylose concentration and c_a the concentration of the dil. solution phase in the phase-separated solution. When the temperature dependence of the rate was treated according to the crystallization of polymers, it was found that the rate was in proportion to T_m²/T(ΔT)², where T_m is the melting point of the polymer in solution and ΔT is (T_m?T). The T_m thus obtained was 120°C for an amylose solution. These results suggested a certain correlation between the amylose retrogradation and the crystallization.     

Hyaluronic Acid Degradation by Ascorbic Acid and Influence of Iron

The effects of ascorbic acid, iron and ADP on hyaluronic acid, a compound present in inflamed joints, were investigated in an in vitro system. Ascorbic acid induces degradation of hyaluronic acid which increased in the presence of FeCl, and which is additionally stimulated by ADP chelated ferric ions. The hyaluronic acid degrading reactions induced by the Fe-III/ADP/ascorbic acid system were inhibited by catalase and formate to various extents whereas the presence of superoxide dismutase did not exert any inhibitory effect. Desferrioxamine, a specific iron chelator, completely inhibited hyaluronic acid depolymerisation by ascorbic acid as well as in combination with FeCl₃ or FeCl₃/ADP, respectively. We suggest that the ultimate hyaluronic acid degrading species is OH', generated via the Fe-III/ADP catalysed Haber Weiss reaction. There is also an indication for the involvement of perferryl or/and ferryl species in the degradation process.     

以L-天冬氨酸为原料制备D-天冬氨酸的新方法

以L-天冬氨酸为原料经过酯化、消旋、拆分和水解制备D-天冬氨酸。使L-2,3-二苯甲酰酒石酸(L-DBTA)与DL-天冬氨酸-β-甲酯在水溶液中于65~70℃反应形成非对映体盐,冷却到室温,D-天冬氨酸.L-DBTA盐析出,过滤后再经水解得到D-天冬氨酸,收率78.2%,旋光纯度达到99%以上。     

氨基酸生产和海洋生物的氨基酸资源开发

氨基酸在医药、食品、饲料等领域有着极为重要和广泛的用途,世界上氨基酸总需求量以５～１０％递增,市场竞争十分激烈。生物资源提取、化学合成、生物合成和综合法是生产氨基酸的４种技术,目前的发展趋势为生物合成和综合法,特别是将现代生物工程技术应用于氨基酸生产。另外,氨基酸生产领域另一个新的倾向是海洋生物氨基酸资源的开发和应用,尤其是海洋生物所产生的特殊氨基酸、肽及其衍生物的开发,同时,综合利用海产品加工后的废弃物来生产氨基酸也受到重视。     

Acid Orcein-Iron and Acid Orcein-Copper Stains for Elastin

Paraffin sections of formol-fixed tissues stained 4-18 hr in 70% alcohol containing 1% orcein and 1% of concentrated (12 N) HCl by volume yield the familiar purple brown elastin and red nuclei on a pink background. When sections so stained are transferred directly from the stain to 70% alcohol containing 0.02% ferric chloride (FeCl₃·6 H₂O) or 0.02% copper sulfate (CuSO₄·5 H₂O) for a 15 sec to 3 min period, elastin coloration is changed to black or reddish black and chromatin staining to reddish black. The procedure can be counterstained with picro-methyl blue to yield blue collagen and reticulum or with our flavianic acid, ferric chloride, acid fuchsin mixture to give deep yellow background and deep red collagen.     

Quinone-Amino Acid Conjugates Targeting Leishmania Amino Acid Transporters

The aim of the present study was to investigate the feasibility of targeting Leishmania transporters via appropriately designed chemical probes. Leishmania donovani, the parasite that causes visceral leishmaniasis, is auxotrophic for arginine and lysine and has specific transporters (LdAAP3 and LdAAP7) to import these nutrients. Probes 1–15 were originated by conjugating cytotoxic quinone fragments (II and III) with amino acids (i.e. arginine and lysine) by means of an amide linkage. The toxicity of the synthesized conjugates against Leishmania extracellular (promastigotes) and intracellular (amastigotes) forms was investigated, as well their inhibition of the relevant amino acid transporters. We observed that some conjugates indeed displayed toxicity against the parasites; in particular, 7 was identified as the most potent derivative (at concentrations of 1 µg/mL and 2.5 µg/mL residual cell viability was reduced to 15% and 48% in promastigotes and amastigotes, respectively). Notably, 6, while retaining the cytotoxic activity of quinone II, displayed no toxicity against mammalian THP1 cells. Transport assays indicated that the novel conjugates inhibited transport activity of lysine, arginine and proline transporters. Furthermore, our analyses suggested that the toxic conjugates might be translocated by the transporters into the cells. The non-toxic probes that inhibited transport competed with the natural substrates for binding to the transporters without being translocated. Thus, it is likely that 6, by exploiting amino acid transporters, can selectively deliver its toxic effects to Leishmania cells. This work provides the first evidence that amino acid transporters of the human pathogen Leishmania might be modulated by small molecules, and warrants their further investigation from drug discovery and chemical biology perspectives.     

Endogenous levels of abscisic Acid and decanoic Acid in dutch iris bulbs and the influence of abscisic Acid and decanoic Acid on iris meristems cultured in vitro

Abscisic acid (ABA) and decanoic acid inhibited shoot elongation and floral development of Dutch iris (Iris hollandica Hoog. cv Ideal) meristems cultured in vitro. No synergism with respect to inhibition of leaf growth between ABA and decanoic acid was observed. With monthly harvest dates, from July 10, 1981 to October 10, 1981, there was a progressive decrease in endogenous level of free ABA in `Ideal' iris bulbs. Bulbs subjected to a full set of the usual preplanting storage conditions flowered, on average, 46 days after planting versus 194 days after planting for bulbs planted directly after harvest. ABA levels at harvest were 4- to 5-fold those after the preplanting storage treatment. In general, ABA levels did not correlate well with the length of time from planting until flowering of iris bulbs. Endogenous decanoic acid levels did not follow any pattern with respect to harvest date or postharvest treatment. After the postharvest high temperature treatment, there was about a 3-fold increase in nonscale decanoic acid concentration. Decanoic acid levels, in nonscale tissue, remained high after each of the other postharvest treatments. It is concluded that there is no good evidence to support the contention that either ABA or decanoic acid is directly involved in iris bulb dormancy.     

Characterization of Excess Deoxyribonucleic Acid Synthesized by Pneumococci in the Presence of Polyadenylic Acid and Deoxyribonucleic Acid Precursors

Excess deoxyribonucleic acid (DNA) synthesized by cell suspensions of encapsulated pneumococci in the presence of polyadenylic acid plus all eight of the naturally occurring deoxyribonucleosides and deoxyribonucleotides has been characterized in several ways. The DNA represents complete molecules, is synthesized by a relatively large population at a steady rate, and is replicated in a semiconservative manner.     

Amino Acid Neurotransmitters in the CNS: Properties of Diaminobutyric Acid Transport

Uptake of L-2,4-diaminobutyric acid (DABA), a positively charged analogue of gamma-aminobutyric acid (GABA), by a synaptosomal fraction isolated from rat brain occurred with a Km of 54 +/- 12 microM and a Vmax of 1.3 +/- 0.2 nmol/min/mg protein. The transport of DABA was inhibited competitively by GABA whereas that of GABA was affected in the same manner by addition of DABA. The maximal accumulation of DABA ([DABA]i/[DABA]c) was observed to increase as the second power of the transmembrane electrical potential ([K+]i/[K+]e) and the first power of the sodium ion concentration gradient. These findings indicate that DABA is transported on the GABA carrier with a net charge of +2, where one charge is provided by the cotransported Na+ and the second is contributed by the amino acid itself. Since uptake of GABA, an electroneutral molecule, is accompanied by transfer of two sodium ions, the results obtained with DABA suggest that one of the sodium binding sites on the GABA transporter is in proximity to the amino acid binding site.     

Enzymatic Formation of Hexadecenoic Acid from Palmitic Acid

Desaturation of palmitic acid was investigated in an enzyme system prepared from rat liver. 2-trans-Hexadecenoic acid as well as 9-cis-hexadecenoic acid (palmitoleic acid) were found to be formed as monoenoic acid in this system.     

Inhibition of Amino Acid Transport in Rabbit Intestine by p-Chloromercuriphenyl Sulfonic Acid

Influx of phenylalanine across the brush border of rabbit intestine is markedly reduced by treatment with 5 mM p-chloromercuriphenyl sulfonate (PCMBS). The effect is rapidly and completely reversed by dithiothreitol. Phenylalanine influx into PCMBS-treated tissue can be competitively inhibited by other neutral amino acids and follows saturation kinetics. PCMBS causes an increase in the apparent Michaelis constant from the value observed in control tissue but does not alter the maximal influx significantly. Treatment of the tissue with PCMBS leads to a significant reduction in the Na-sensitivity of the transport, and a number of results indicate that the major effect of the reagent is to cause a marked reduction in the affinity of the transport system for Na. The transport system can be partially protected against reaction with PCMBS by phenylalanine and tryptophan but not by methionine or norleucine. The results suggest that PCMBS reacts with a sulfhydryl group in the region of the transport site and may alter conformational changes associated with the binding of substrates.     

Engineering Filamentous Fungi for Conversion of d-Galacturonic Acid to l-Galactonic Acid

d-Galacturonic acid, the main monomer of pectin, is an attractive substrate for bioconversions, since pectin-rich biomass is abundantly available and pectin is easily hydrolyzed. l-Galactonic acid is an intermediate in the eukaryotic pathway for d-galacturonic acid catabolism, but extracellular accumulation of l-galactonic acid has not been reported. By deleting the gene encoding l-galactonic acid dehydratase (lgd1 or gaaB) in two filamentous fungi, strains were obtained that converted d-galacturonic acid to l-galactonic acid. Both Trichoderma reesei Δlgd1 and Aspergillus niger ΔgaaB strains produced l-galactonate at yields of 0.6 to 0.9 g per g of substrate consumed. Although T. reesei Δlgd1 could produce l-galactonate at pH 5.5, a lower pH was necessary for A. niger ΔgaaB. Provision of a cosubstrate improved the production rate and titer in both strains. Intracellular accumulation of l-galactonate (40 to 70 mg g biomass⁻¹) suggested that export may be limiting. Deletion of the l-galactonate dehydratase from A. niger was found to delay induction of d-galacturonate reductase and overexpression of the reductase improved initial production rates. Deletion of the l-galactonate dehydratase from A. niger also delayed or prevented induction of the putative d-galacturonate transporter An14g04280. In addition, A. niger ΔgaaB produced l-galactonate from polygalacturonate as efficiently as from the monomer.     

Stereoisomeric Characterization of Tartaric Acid Produced during l-Ascorbic Acid Metabolism in Plants

Labeled tartaric acids from Pelargonium crispum apices which had been fed l-ascorbic acid-6-(14)C and Vitis labrusca and Parthenocissus inserta tissues which had been fed l-ascorbic acid-1-(14)C were examined by chemical means to determine chiral configuration. In each instance, label was associated with (+)-tartaric acid.Similar experiments with labeled tartaric acid from P. crispum which had been labeled with d-glucose-1-(14)C or -6-(14)C led to the same result. No evidence was obtained for formation of labeled meso-tartaric acid in experiments described above. The recent suggestion of H. Ruffner and D. Rast (Z. Pflanzenphysiol. 73: 45-55, 1974) that conversion of l-ascorbic acid to tartaric acid in plants is a nonenzymatic process is re-examined in the light of present findings.