Loss of aldehyde dehydrogenase in an Escherichia coli mutant selected for growth on the rare sugar L-galactose.

Escherichia coli K-12 converts L-fucose to dihydroxyacetone phosphate (C-1 to C-3) and L-lactaldehyde (C-4 to C-6) by a pathway specified by the fuc regulon. Aerobically, L-lactaldehyde serves as a carbon and energy source by the action of an aldehyde dehydrogenase of broad specificity; the product, L-lactate, is then converted to pyruvate. Anaerobically, L-lactaldehyde serves as an electron acceptor to regenerate NAD from NADH by the action of an oxidoreductase; the reduced product, L-12-propanediol, is excreted. A strain selected for growth on L-galactose (a structural analog of L-fucose) acquired a broadened inducer specificity because of an altered fucR gene encoding the activator protein for the fuc regulon (Y. Zhu and E. C. C. Lin, J. Mol. Evol. 23:259-266, 1986). In this study, a second mutation that abolished aldehyde dehydrogenase activity was discovered. The L-fucose pathway converts L-galactose to dihydroxyacetone phosphate and L-glyceraldehyde. Aldehyde dehydrogenase then converts L-glyceraldehyde to L-glycerate, which is toxic. Loss of the dehydrogenase averts the toxicity during growth on L-galactose, but reduces by one-half the aerobic growth yield on L-fucose. When mutant cells induced in the L-fucose system were incubated with radioactive L-fucose, accumulation of radioactivity occurred if the substrate was labeled at C-1 but not if it was labeled C-6. Complete aerobic utilization of carbons 4 through 6 of L-fucose depends not only on an adequate activity of aldehyde dehydrogenase to trap L-lactaldehyde as its anionic acid but also on the lack of L-1,2-propanediol oxidoreductase activity, which converts L-lactaldehyde to a readily excreted alcohol.     

A comparison of alternate metabolic strategies for the utilization of D-arabinose

Summary Mutants ofKlebsiella aerogenes W70 that metabolize the uncommon pentose D-arabinose were isolated. These mutants were found to be either constitutive or indicible by D-arabinose for the synthesis of enzymes in the L-fucose pathway. Such mutants could then utilize L-fucose isomerase to convert the structurally similar D-arabinose molecule to D-ribulose. D-Ribulose is an inter-mediate and the inducer of an existing ribitol pathway and could thus be metabolized. In those D-arabinose-positive mutants where the ribitol pathway was blocked by mutation, D-ribulose could alternatively be metabolized by using the remaining L-fucose pathway enzymes. When the two D-arabinose catabolic routes were compared, catabolism of D-arabinose via the ribitol pathway was found to be more efficient. Catabolism of D-arabinose using the L-fucose pathway per-mitted D-ribulose to escape into the media and produced an unmetabolizable end product, L-glycolic acid. A comparison of growth using constitutive versus inducible control of the borrowed L-fucose isomerase did not reveal an advantage for one control type over the other. Several differences were observed,however, when we determined the degree to which these control mutations perturbed the normal functioning of the L-fucose and associated pathways. Growth of the constitutive mutant was impaired with L-fucose as substrate. The inducible-control mutant had altered growth characteristics on ribitol and L-rhamnose.     

Binding of different monosaccharides by lectin PA-IIL from Pseudomonas aeruginosa: thermodynamics data correlated with X-ray structures

The lectin from Pseudomonas aeruginosa (PA-IIL) is involved in host recognition and biofilm formation. Lectin not only displays an unusually high affinity for fucose but also binds to L-fucose, L-galactose and D-arabinose that differ only by the group at position 5 of the sugar ring. Isothermal calorimetry experiments provided precise determination of affinity for the three methyl-glycosides and revealed a large enthalpy contribution. The crystal structures of the complexes of PA-IIL with L-galactose and Met-beta-D-arabinoside have been determined and compared with the PA-IIL/fucose complex described previously. A combination of the structures and thermodynamics provided clues for the role of the hydrophobic group in affinity.     

Impact of oxidative stress on ascorbate biosynthesis in Chlamydomonas via regulation of the VTC2 gene encoding a GDP-L-galactose phosphorylase

The L-galactose (Smirnoff-Wheeler) pathway represents the major route to L-ascorbic acid (vitamin C) biosynthesis in higher plants. Arabidopsis thaliana VTC2 and its paralogue VTC5 function as GDP-L-galactose phosphorylases converting GDP-L-galactose to L-galactose-1-P, thus catalyzing the first committed step in the biosynthesis of L-ascorbate. Here we report that the L-galactose pathway of ascorbate biosynthesis described in higher plants is conserved in green algae. The Chlamydomonas reinhardtii genome encodes all the enzymes required for vitamin C biosynthesis via the L-galactose pathway. We have characterized recombinant C. reinhardtii VTC2 as an active GDP-L-galactose phosphorylase. C. reinhardtii cells exposed to oxidative stress show increased VTC2 mRNA and L-ascorbate levels. Genes encoding enzymatic components of the ascorbate-glutathione system (e.g. ascorbate peroxidase, manganese superoxide dismutase, and dehydroascorbate reductase) are also up-regulated in response to increased oxidative stress. These results indicate that C. reinhardtii VTC2, like its plant homologs, is a highly regulated enzyme in ascorbate biosynthesis in green algae and that, together with the ascorbate recycling system, the L-galactose pathway represents the major route for providing protective levels of ascorbate in oxidatively stressed algal cells.     

L-Ascorbate biosynthesis in higher plants: the role of VTC2

In the past year, the last missing enzyme of the L-galactose pathway, the linear form of which appears to represent the major biosynthetic route to L-ascorbate (vitamin C) in higher plants, has been identified as a GDP-L-galactose phosphorylase. This enzyme catalyzes the first committed step in the synthesis of that vital antioxidant and enzyme cofactor. Here, we discuss how GDP-L-galactose phosphorylase enzymes, encoded in Arabidopsis by the paralogous VTC2 and VTC5 genes, function in concert with the other enzymes of the L-galactose pathway to provide plants with the appropriate levels of L-ascorbate. We hypothesize that regulation of L-ascorbate biosynthesis might occur at more than one step and warrants further investigation to allow for the manipulation of vitamin C levels in plants.     

Natural and altered induction of the L-fucose catabolic enzymes in Klebsiella aerogenes.

Mutants of Klebsiella aerogenes W70 were isolated that had gained the ability to utilize the uncommon pentose D-arabinose as their sole source of carbon and energy. In contrast to the D-arabinose-negative, parent strain, these mutants were found to be either constitutive for certain enzymes of the L-fucose catabolic pathway or inducible for such enzymes when incubated in the presence of D-arabinose. The mutants used L-fucose isomerase to convert D-arabinose to D-ribulose, which is an intermediate and inducer of the ribitol catabolic pathway. The D-ribulokinase of the ribitol pathway was then induced. This enzyme catalyzed the phosphorylation of D-ribulose at the 5-carbon position. Mutants that were negative for D-ribulokinase could still dissimilate D-arabinose slowly by using all three enzymes, the isomerase, kinase, and aldolase, of the L-fucose pathway. Using condition negative mutants, we were able to demonstrate that the natural induction of the L-fucose pathway enzymes by L-fucose required the activity of a functional L-fucose isomerase and a functional L-fuculokinase but not an L-fuculose-1-phosphate aldolase. A metabolic intermediate, L-fuculose-1-phosphate, was thereby shown to be a probable inducer of at least the isomerase and kinase of the L-fucose catabolic pathway. Similar experiments, with D-arabinose-positive mutants, which were induced for the L-fucose pathway enzymes upon incubation with D-arabinose, revealed that the activities of the L-fucose isomerase and the L-fuculokinase were also required for the induction of the L-fucose enzymes. These D-arabinose-positive mutants apparently produced an altered regulatory protein that accepted both L-fuculose-1-phosphate and D-ribulose-1-phosphate as inducers. Examination of constitutive mutants revealed that L-fucose isomerase and L-fuculokinase were both synthesized constitutively, with the aldolase apparently under separate control.     

Construction of an improved D-arabinose pathway in Escherichia coli K-12.

A ribitol catabolic pathway was transduced into Escherichia coli K-12 in an effort to determine whether the ribitol pathway would confer an advantage to D-arabinose-positive mutants growing on D-arabinose as the sole carbon source. Competition studies in chemostats showed that ribitol-positive strains, with a selection coefficient of 9%/h, have a significant competitive advantage over ribitol-negative strains. Ribitol-positive strains grown in batch culture also exhibited a shorter lag period than did ribitol-negative strains when transferred from glucose to D-arabinose. Repeated transfer of a ribitol-positive strain of E. coli K-12 on D-arabinose yielded a strain with further improved growth on D-arabinose. This "evolved" strain was found to constitutively synthesize L-fucose permease, isomerase, and kinase but had lost the ability to grow on L-fucose, apparently owing to the loss of a functional aldolase. This constitutive mutation is not linked to the fucose gene cluster and may be similar to an unlinked constitutive mutation described by Chen et al. (J. Bacteriol. 159:725-729, 1984).     

Molecular cloning of the Escherichia coli B L-fucose-D-arabinose gene cluster.

To metabolize the uncommon pentose D-arabinose, enteric bacteria often recruit the enzymes of the L-fucose pathway by a regulatory mutation. However, Escherichia coli B can grow on D-arabinose without the requirement of a mutation, using some of the L-fucose enzymes and a D-ribulokinase that is distinct from the L-fuculokinase of the L-fucose pathway. To study this naturally occurring D-arabinose pathway, we cloned and partially characterized the E. coli B L-fucose-D-arabinose gene cluster and compared it with the L-fucose gene cluster of E. coli K-12. The order of the fucA, -P, -I, and -K genes was the same in the two E. coli strains. However, the E. coli B gene cluster contained a 5.2-kb segment located between the fucA and fucP genes that was not present in E. coli K-12. This segment carried the darK gene, which encodes the D-ribulokinase needed for growth on D-arabinose by E. coli B. The darK gene was not homologous with any of the L-fucose genes or with chromosomal DNA from other D-arabinose-utilizing bacteria. D-Ribulokinase and L-fuculokinase were purified to apparent homogeneity and partially characterized. The molecular weights, substrate specificities, and kinetic parameters of these two enzymes were very dissimilar, which together with DNA hybridization analysis, suggested that these enzymes are not related. D-Arabinose metabolism by E. coli B appears to be the result of acquisitive evolution, but the source of the darK gene has not been determined.     

Isolation and characterization of L-fucose dehydrogenase from rabbit liver

L-Fucose dehydrogenase [EC 1.1.1.122] was isolated from a rabbit liver extract and purified about 390-fold with a yield of approximately 13%. The purification procedures included treatment with protamine, ammonium sulfate fractionation, treatment with acid, DE-32 celluose colum chromatography, gel filtration on Sephadex G-100, preparative polyacrylamide gel electrophoresis, and affinity chromatography on 5' AMP-Sepharose 4B. The last procedure, affinity chromatography on 5' AMP-Sephadex 4B, was useful for the removal of other dehydrogenases. The eznyme which was homogeneous, as shown by polyacrylamide gel electrophoresis, had a molecular weight of about 92,000. The optimum pH was at 10.0 and isoelectric point at 5.2. The enzyme accepted both L-fucose and D-arabinose as substrate, but was specific for NAD+ as coenzyme. Km values were 0.15 mM, 1.4 mM, and 0.7 mM for L-fucose, D-arabinose, and NAD+, respectively. A single enzyme catalyzed the oxidation of L-fucose and D-arabinose, which had the same configurations of hydroxyl groups from C-2 to C-4. The reaction products obtained with L-fucose as substrate were L-fucono-lactone and L-fuconic acid. The L-fucono-lactone was an immediate product of oxidation and was hydrolyzed to L-fuconic acid spontaneously. This reaction was irreversible. Therefore, it is likely that L-fucose dehydrogenase is involved in the initial step of the catabolic pathway of L-fucose in rabbit liver.     

Identification of a salvage pathway for D-arabinose in Mycobacterium smegmatis

Extracts of Mycobacterium smegmatis, which was adapted to growth in synthetic medium containing D-arabinose as sole carbon source, catalyzed the NADPH-mediated reduction of D-arabinose to D-arabitol. When arabinose-adapted bacteria were transferred to glycerol medium, resumption of growth was accompanied by a sharp drop in the specific activity of this enzyme. Moreover, extracts of cells grown in D-arabinose medium contained large amounts of an NAD+-linked pentitol dehydrogenase, as compared to bacteria multiplying in glycerol medium. The specific activity of mycobacterial extracts was ten-fold higher for D-arabitol than for its L-isomer, and eight-fold higher than for xylitol (it was more than forty-fold lower in the case of glycerol-grown cells). The product of the pentitol dehydrogenase reaction was identified as D-xylulose by three different procedures. On the basis of these data, it is suggested that utilization of exogenous D-arabinose in mycobacteria involves two dehydrogenases that catalyze the reactions D-arabinose NADPH----D-arabitol NAD+----D-xylulose, by virtue of which an aldopentose is converted into a ketopentose. The alditol: NADP oxidoreductase was isolated from homogenates of D-arabinose-adapted mycobacteria, and purified by DEAE-cellulose chromatography. The enzymatic activity was restricted to a single band which, under denaturing conditions, comigrated with albumin (approximately 46 kDa). It was insensitive to 2-mercaptoethanol, EDTA and NaF, and was inactivated at 70 degrees C.     

D-arabinose metabolism in Escherichia coli B: induction and cotransductional mapping of the L-fucose-D-arabinose pathway enzymes.

D-Arabinose is degraded by Escherichia coli B via some of the L-fucose pathway enzymes and a D-ribulokinase which is distinct from the L-fuculokinase of the L-fucose pathway. We found that L-fucose and D-arabinose acted as the apparent inducers of the enzymes needed for their degradation. These enzymes, including D-ribulokinase, appeared to be coordinately regulated, and mutants which constitutively synthesized the L-fucose enzymes also constitutively synthesized D-ribulokinase. In contrast to D-arabinose-positive mutants of E. coli K-12, in which L-fuculose-1-phosphate and D-ribulose-1-phosphate act as inducers of the L-fucose pathway, we found that these intermediates did not act as inducers in E. coli B. To further characterize the E. coli B system, some of the L-fucose-D-arabinose genes were mapped by using bacteriophage P1 transduction. A transposon Tn10 insertion near the E. coli B L-fucose regulon was used in two- and three-factor reciprocal crosses. The gene encoding D-ribulokinase, designated darK, was found to map within the L-fucose regulon, and the partial gene order was found to be Tn10-fucA-darK-fucI-fucK-thyA.     

Production,properties and specificity of a new bacterial L-fucose- and D-arabinose-binding lectin of the plant aggressive pathogen Ralstonia solanacearum,and its comparison to related plant and microbial lectins

The worldwide distributed plant aggressive pathogen Ralstonia solanacearum, which causes lethal wilt in many agricultural crops, produces a potent L-fucose-binding lectin (RSL) exhibiting sugar specificity similar to that of PA-IIL of the human aggressive opportunistic pathogen Pseudomonas aeruginosa. Both lectins show L-fucose > L-galactose > D-arabinose > D-mannose specificity, but the affinities of RSL to these sugars are substantially lower. Unlike Ulex europaeus anti-H lectin, but like PA-IIL and Aleuria aurantia lectin (AAL), RSL agglutinates H-positive human erythrocytes regardless of their type, O, A, B, or AB, and animal erythrocytes (papain-treated ones more strongly than untreated ones). It also interacts with H and Lewis chains in the saliva of "secretors" and "nonsecretors." RSL purification is easier than that of PA-IIL since R. solanacearum extracts do not contain a galactophilic PA-IL-like activity. Mass spectrometry and 35 N-terminal amino acid sequencing enabled identification of the RSL protein (subunit approximately 9.9 kDa, approximately 90 amino acids) in the complete genome sequence of this bacterium. Despite the greater phylogenetic proximity of R. solanacearum to P. aeruginosa, and the presence of a PA-IIL-like gene in its genome, the RSL structure is not related to that of PA-IIL, but to that of the fucose-binding lectin of the mushroom (fungus) Aleuria aurantia, which like the two bacteria is a soil inhabitant.     

Oxidation of L-glucose by a Pseudomonad.

A new enzyme, D-threo-aldolse dehydrogenase (2S,3R-aldose dehydrogenase), found in Pseudomonas caryophylli, was capable of oxidizing L-glucose L-xylose, D-arabinose, and L-fucose in the presence of NAD+. The enzyme was synthesized constitutively and purified about 120-fold from D-glucose-grown cells. The Km values for L-glucose, L-xylose, D-arabinose, and L-fucose were 1.5 . 10(-2), 4.5 . 10(-3), 2.8 . 10(-3), and 2.1 . 10(-3), respectively. D-glucose and other aldoses inhibited the enzyme reaction; this inhibition was competitive with L-glucose as substrate and D-glucose as inhibitor. The optimum pH for the enzyme reaction was 10; the molecular weight of the enzyme was determined by gel filtration to be 7 . 10(4).     

Note on study of the sporulation of fungi: endotrophic sporulation in the genusPenicillium

Conidiophore formation and sporulation can be induced inPenicillium sp. strain P 17 by an environmental factor—carbohydrate (carbon) starvation. Both surface and submerged mycelium, when transferred from synthetic medium to glucose-free salt solution, form conidiophores and sporulate, while in the control cultures on complete medium, vegetative growth continues. The time required for the formation of conidiophores, i.e. the induction interval, is 7–14 h and its length increases with the age of both surface and submerged mycelia. During the induction phase the mycelium undergoes autolysis, associated with degradation of energy motabolism involving the comsumption of reserve substances, a rapid drop in endogenous respiration and the endogenous reducing activity of the mycelium, a decrease in the labile phosphate concentration, proteolysis, an increase in the ammonia and orthopsphate concentration and exhaustion of readily oxidized amino acids from the pool. A transient increase in respiration occurs before differentiation of the conidiophores starts. During the second half of the induction phase, polyphenol substances and polyphenol oxidase appear in the mycelium.The enzyme is not induced by exogenous phenols. Its possible role in the sporulation of fungi is considered.     

Proton-linked L-fucose transport in Escherichia coli.

1. Addition of L-fucose to energy-depleted anaerobic suspensions of Escherichia coli elicited an uncoupler-sensitive alkaline pH change diagnostic of L-fucose/H+ symport activity. 2. L-Galactose or D-arabinose were also substrates, but not inducers, for the L-fucose/H+ symporter. 3. L-Fucose transport into subcellular vesicles was dependent upon respiration, displayed a pH optimum of about 5.5, and was inhibited by protonophores and ionophores. 4. These results showed that L-fucose transport into E. coli was energized by the transmembrane electrochemical gradient of protons. 5. Neither steady state kinetic measurements nor assays of L-fucose binding to periplasmic proteins revealed the existence of a second L-fucose transport system.     

Hydrophobic-ionic chromatography. Its application to purification of porcine pancreas enzymes.

1. All the porcine pancreas enzymes tested, regardless of their pI's were adsorbed on Amberlite CG-50 (a weakly acidic cation exchange resin) at pH 4, where the ion-exchange group (carboxyl group) is not dissociated. The adsorption is hardly influenced by ionic strength. 2. At pH 4, the adsorbed enzymes were partially eluted by organic solvents such as 50% propanol. 3. The adsorbed enzymes were effectively eluted by increasing the pH from 4 to 6. Trypsin (pI 10.5) was eluted before carboxypeptidase A (pI 4.5 AND 5.3) WITH 0.5 M acetate buffer, whereas the former enzyme was eluted after the latter enzyme with 0.2 M 3,3-dimethyl glutarate buffer. However, with either buffer, the elution order of enzymes was not always the same as the order of the pI's. 4. By a single Amberlite CG-50 column chromatography of porcine pancreas extracts, kallikrein, carboxypeptidase B, deoxyribonuclease, carboxypeptidase A, and trypsin were purified 100-fold, 16-fmately 13%. The purification procedures included treatment with protamine, ammonium sulfate fractionation, treatment with acid, DE-32 cellulose column chromatography, gel filtration on Sephadex G-100, preparative polyacrylamide gel electrophoresis, and affinity chromatography on 5' AMP-Sepharose 4B. The last procedure, affinity chromatography on 5' AMP-Sepharose 4B, was useful for the removal of other dehydrogenases. The enzyme which was homogeneous, as shown by polyacrylamide gel electrophoresis, had a molecular weight of about 92,000. The optimum pH was at 10.0 and isoelectric point at 5.2. The enzyme accepted both L-fucose and D-arabinose as substrate, but was specific for NAD+ as coenzyme. Km values were 0.15 mM, 1.4 mM, and 0.07 mM for L-fucose, D-arabinose, and NAD+, respectively. A single enzyme catalyzed the oxidation of L-fucose and D-arabinose, which had the same configurations of hydroxyl groups from C-2 to C-4. The reaction products obtained with L-fucose as substrate were L-fucono-lactone and L-fuconic acid. The L-fucono-lactone was an immediate product of oxidation and was hydrolyzed to L-fuconic acid spontaneously. This reaction was irreversible. Therefore, it is likely that L-fucose dehydrogenase is involved in the initial step of the catabolic pathway of L-fucose in rabbit liver.     

Estrogenic reduction of styrene monomer degraded by Phanerochaete chrysosporium KFRI 20742

The characteristic biodegradation of monomeric styrene by Phanerochaete chrysosporium KFRI 20742, Trametes versicolor KFRI 20251 and Daldinia concentrica KFRI 40-1 was carried out to examine the resistance, its degradation efficiency and metabolites analysis. The estrogenic reduction effect of styrene by the fungi was also evaluated. The mycelium growth of fungi differentiated depending on the concentration levels of styrene. Additionally P. chrysosporium KFRI 20742 showed superior mycelium growth at less than 200 mg/l, while D. concentrica KFRI 40-1 was more than 200 mg/l. The degradation efficiency reached 99% during one day of incubation for all the fungi. Both manganese-dependent peroxidase and laccase activities in liquid medium were the highest at the initial stage of incubation, whereas the lowest was after the addition of styrene. However, both activities were gradually recovered after. The major metabolites of styrene by P. chrysosporium KFRI 20742 were 2-phenyl ethanol, benzoic acid, cyclohexadiene-1,4-dione, butanol and succinic acid. From one to seven days of incubating the fungi, the expression of pS2 mRNA widely known as an estrogen response gene was decreased down to the level of baseline after one day. Also, the estrogenic effect of styrene completely disappeared after treatment with supernatant of P. chrysosporium KFRI 20742 from one week of culture down to the levels of vehicle.     

Role of uronic acids present in phytopathogenic fungi as inducers of polygalacturonases during autolysis

The presence of uronic acids in the culture fluid and mycelium of the fungi: Alternaria alternata, Botrytis cinerea, Drechslera halodes, Fusarium culmorum, Fusarium oxysporum, Monilinia fructigena, Mucor mucedo, Rhizopus stolonifer and Trichoderma hamatum was detected and quantified. In these fungi the concentration of uronic acids increased during the growth phase and the maximal concentrations were found at the end of the growth phase or onset of autolysis both in the mycelium as well as in the culture fluid. The uronic acids were metabolized during the first days of autolysis decreasing to constant levels until the end of the autolytic period studied.The variations in the activity of polygalacturonase and polymethylgalacturonase present in the culture fluid were determined at the onset and during autolysis in these fungi. These enzymic activities were found in the culture fluid of these fungi, with exception of M. rouxii, and they showed an increasing activity in the first days of autolysis and later a slight increase or decrease was observed. The presence of uronic acids in these phytopathogenic or saprophytic fungi and the low levels detected during autolysis could be related to the induction of pectic enzymes and the pathogenicity of these fungi.     

L-fucose metabolism in mammals. Kinetic studies on pork liver 2-keto-3-deoxy-L-fuconate:NAD+ oxidoreductase.

Pork liver 2-keto-3-deoxy-L-fuconate:NAD+ oxidoreductase has been shown to convert 2-keto-3-deoxy-L-fuconate to a 6-carbon acid tentatively identified as 2,4(or 5)-diketo-5(or 4)-monohydroxyhexanoate. The enzyme has a pH optimum of 10. 5 or higher. It is stabilized by dithiothereitol and inhibited by p-hydroxymercuribenzoate and heavy metals (Ag+, Hg2+, Co2+, Cd2+, Pb2+, Zn2+, and Cu2+), suggesting the presence of a functionally essential sulfhydryl group; pre-treatment of enzyme with NAD+ prevents inhibition by p-hydrocymercuribenzoate and heavy metals indicating that this sulfhydryl group may be near the NAD+ binding site. The enzyme has an absolute requirement for NAD+; NADP+ is an ineffective coenzyme. Several lines of evidence indicate that the same enzyme acts on both 2-keto-3-deocy-L-fuconate and 2-keto-3-deoxy-D-arabonate; thus, the pure enzyme acts on both substrates, the two substrates have very similar kinetic parameters (Km values are: 2-keto-3-deocy-L-fuconate, 0.20 mM; 2-keto-3-deoxy-D-arabonate, 0.25 mM; NAD+ for either substrate, 0.22 to 0.25 mM), the two substrates show identical pH and temperature profiles and the two substrates compete for common enzyme active sites. A large number of other sugars and sugar acids, including several 2-keto-3-deoxyaldonates, were ineffective as substrates. The dehydrogenase was also found in calf, beef, lamb, mouse, and rat liver. These studies when considered together with previous studies on the metabolism of L-fucose in pork liver indicate the presence of a soluble enzyme pathway capable of converting L-fucose to 2,4(or 5)-diketo-5(or 4)-monohydroxyhexanoate; this pathway can also convert D-arabinose, and probably L-galactose, to the analogous derivatives (diketomonohydroxypentanoate and diketodihydroxyhexanoate, respectively.     

温度对外源性^32P在水、铜绿微囊藻和底泥中迁移的影响

采用同位素示踪法,在实验室模拟研究不同温度下外源性无机磷酸盐在水、铜绿微囊藻(Micro-fystis aeruginoas)和底泥中的迁移过程,外源性³²P加入水中后,首先是一种与温度无关的快速物理化学分配,大量溶解性磷酸盐迅速进入底泥和微囊藻中,随后水中³²P的迁移主要受微囊藻生长状况的影响,温度升高有利于微囊藻的生长,并提高了微囊藻吸磷的速度,微囊藻中最大外源性磷浓度只与水环境中的初始磷浓度有关,25℃时铜绿微囊藻的生长曲线有7d的对数期,没有明显的稳定期就转入衰亡期,在25℃时,当微囊藻超积累P到一定程度后,其对数生长同细胞内含P量无关,随着时间的推移,外源性³²P不断向底泥中迁移,实验末期所有的³²P都转移到底泥中,提高温度使水中溶解性外源性磷的下降速率加快,7d后水中溶解的外源性磷浓度低于0．00716mg·L^-1。