ABSTRACT
Bioactive natural C-glycosides are rare and chemical C-glycosylation faces challenges while enzymatic C-glycosylation catalyzed by C-glycosyltransferases provides an alternative way. However, only a small number of C-glycosyltransferases have been found, and most of the discovered C-glycosyltransferases prefer to glycosylate phenols with an acyl side chain. Here, a promiscuous C-glycosyltransferase, AbCGT, which is capable of C-glycosylating scaffolds lacking acyl groups, is identified from Aloe barbadensis. Based on the substrate promiscuity of AbCGT, 16 C-glycosides with inhibitory activity against sodium-dependent glucose transporters 2 are chemo-enzymatically synthesized. The C-glycoside 46a shows hypoglycemic activity in diabetic mice and is biosynthesized with a cumulative yield on the 3.95 g Lâ1 scale. In addition, the key residues involved in the catalytic selectivity of AbCGT are explored. These findings suggest that AbCGT is a powerful tool in the synthesis of lead compounds for drug discovery and an example for engineering the catalytic selectivity of C-glycosyltransferases.
Subject(s)
Aloe/enzymology , Glycosides/biosynthesis , Glycosyltransferases/metabolism , Plant Proteins/metabolism , Sodium-Glucose Transporter 2 Inhibitors/metabolism , Alloxan/toxicity , Aloe/genetics , Animals , Biocatalysis , Blood Glucose/analysis , Blood Glucose/drug effects , Cloning, Molecular , Diabetes Mellitus, Experimental/blood , Diabetes Mellitus, Experimental/chemically induced , Diabetes Mellitus, Experimental/drug therapy , Diabetes Mellitus, Type 1/blood , Diabetes Mellitus, Type 1/chemically induced , Diabetes Mellitus, Type 1/drug therapy , Drug Discovery/methods , Female , Glycosides/pharmacology , Glycosides/therapeutic use , Glycosylation , Glycosyltransferases/genetics , Glycosyltransferases/isolation & purification , Humans , Male , Mice , Plant Proteins/genetics , Plant Proteins/isolation & purification , Recombinant Proteins/genetics , Recombinant Proteins/isolation & purification , Recombinant Proteins/metabolism , Sodium-Glucose Transporter 2/metabolism , Sodium-Glucose Transporter 2 Inhibitors/pharmacology , Sodium-Glucose Transporter 2 Inhibitors/therapeutic use , Substrate SpecificityABSTRACT
The main polysaccharide of the gel present in the leaves of or Aloe vera Burm.F., (Aloe barbadensis Miller) a xerophytic crassulacean acid metabolism (CAM) plant, is an acetylated glucomannan named acemannan. This polysaccharide is responsible for the succulence of the plant, helping it to retain water. In this study we determined using polysaccharide analysis by carbohydrate gel electrophoresis (PACE) that the acemannan is a glucomannan without galactose side branches. We also investigated the expression of the gene responsible for acemannan backbone synthesis, encoding a glucomannan mannosyltransferase (GMMT, EC 2.4.1.32), since there are no previous reports on GMMT expression under water stress in general and specifically in Aloe vera. It was found by in silico analyses that the GMMT gene belongs to the cellulose synthase-like A type-9 (CSLA9) subfamily. Using RT-qPCR it was found that the expression of GMMT increased significantly in Aloe vera plants subjected to water stress. This expression correlates with an increase of endogenous ABA levels, suggesting that the gene expression could be regulated by ABA. To corroborate this hypothesis, exogenous ABA was applied to non-water-stressed plants, resulting in a significant increase of GMMT expression after 48â¯h of ABA treatment.
Subject(s)
Abscisic Acid/pharmacology , Aloe/genetics , Gene Expression Regulation, Enzymologic/drug effects , Gene Expression Regulation, Plant/drug effects , Genes, Plant , Mannans/metabolism , Methyltransferases/genetics , Stress, Physiological , Water/metabolism , Aloe/enzymology , Aloe/metabolism , Amino Acid Sequence , Base Sequence , Catalytic Domain , DNA, Complementary/genetics , Droughts , Electrophoresis, Starch Gel/methods , Gas Chromatography-Mass Spectrometry , Methyltransferases/chemistry , Methyltransferases/metabolism , Sequence Homology, Amino AcidABSTRACT
The glycosides of 4'-demethylepipodophyllotoxin (DMEP) possess various pharmacological activities; however, the chemical synthesis of these glycosides faces challenges in regioselectivity, stereoselectivity, and the protection and de-protection of functional groups. In this work, a novel glycosyltransferase (GT) gene AbGT5 from Aloe barbadensis was successfully cloned, heterogeneously expressed and purified. Recombinant AbGT5 was able to catalyze the glycosylation of DMEP and the glycosylated product, which was separated from the preparative scale reaction, was characterized as DMEP 4'-O-ß-D-glucoside via MS, 1H-NMR, 13C-NMR, HSQC and HMBC. According to the investigations of enzyme properties, AbGT5 show the highest activity around 20 â in the buffer of pH 9.0, and it was independent of divalent metal ions. Under the optimum conditions, the conversion rate of DMEP can reach 80%. Above all, in this work the enzymatic glycosylation of DMEP was achieved with high efficiency by the novel GT AbGT5.
Subject(s)
Glucosides/chemistry , Glycosides/chemistry , Glycosyltransferases/metabolism , Podophyllotoxin/analogs & derivatives , Aloe/enzymology , Aloe/genetics , Glycosylation , Glycosyltransferases/genetics , Podophyllotoxin/chemistryABSTRACT
Acetylated polymannan polysaccharide (ApmP) isolated from Aloe barbadensis Miller contains a stable peroxidase that was solubilized to investigate its biochemical, electrophoretic, immunological, and proteomic properties. In the electrophoretic band corresponding to the solubilized peroxidase, proteomic analysis detected seven tryptic peptides that matched homologous peptides covering one third of the ATP22a peroxidase of Arabidopsis thaliana. All the characteristics tested indicated that the activity stabilized within the ApmP pertains to the basic secretory peroxidase family, which includes members that have several biotechnological uses. Hence ApmP might yield a widely used peroxidase in stabilized form.
Subject(s)
Aloe/enzymology , Mannans/chemistry , Peroxidases/isolation & purification , Plant Extracts/chemistry , Plant Proteins/isolation & purification , Acetylation , Aloe/chemistry , Amino Acid Sequence , Arabidopsis/enzymology , Arabidopsis/genetics , Benzidines/chemistry , Electrophoresis, Polyacrylamide Gel , Hydrogen Peroxide/chemistry , Kinetics , Mass Spectrometry , Molecular Sequence Data , Peptides/analysis , Peroxidases/genetics , Peroxidases/metabolism , Plant Proteins/genetics , Plant Proteins/metabolism , Proteomics , Solubility , TrypsinABSTRACT
Leaf exudates from Aloe species, such as the Southern African Aloe ferox, are used in traditional medicines for both humans and livestock. This includes aloesin, a skin bleaching product that inhibits the synthesis of melanin. Aloesin, (a C-glycoside-5-methylchromone) can be released from aloeresin A, an ester of aloesin, through hydrolysis. The objective of the current study was to identify an enzymatic hydrolysis method for converting aloeresin A to aloesin, resulting in increased concentrations of aloesin in the aloe bitters extract. More than 70 commercially available hydrolytic enzymes were screened for the conversion of aloeresin A. An esterase (ESL001-02) from Diversa, a lipase (Novozym 388) and a protease (Aspergillus oryzae) preparation were identified during screening as being capable of providing conversion of pure aloeresin A, with the protease giving the best conversion (~100%). It was found that a contaminating enzyme in Novo 388 was responsible for the conversion of aloeresin A to aloesin. This contaminating enzyme, possibly a protease, was able to give almost complete conversion using crude aloe bitters extract, doubling the concentration of aloesin in aloe bitters extract via the hydrolysis of aloeresin A.
Subject(s)
Aloe/chemistry , Biocatalysis , Chromones/isolation & purification , Chromones/metabolism , Glucosides/isolation & purification , Glucosides/metabolism , Skin Lightening Preparations/isolation & purification , Africa, Southern , Aloe/enzymology , Aspergillus oryzae/enzymology , Chromones/chemistry , Esterases/metabolism , Glucosides/chemistry , Hydrolysis , Lipase/metabolism , Peptide Hydrolases/metabolism , Plant Extracts/chemistry , Plant Leaves/chemistry , Plant Leaves/enzymology , Skin Lightening Preparations/chemistry , Skin Lightening Preparations/metabolismABSTRACT
Aloe arborescens is a medicinal plant rich in aromatic polyketides, such as pharmaceutically important aloenin (hexaketide), aloesin (heptaketide) and barbaloin (octaketide). Three novel type III polyketide synthases (PKS3, PKS4 and PKS5) were cloned and sequenced from the aloe plant by cDNA library screening. The enzymes share 85-96% amino acid sequence identity with the previously reported pentaketide chromone synthase and octaketide synthase. Recombinant PKS4 and PKS5 expressed in Escherichia coli were functionally identical to octaketide synthase, catalyzing the sequential condensations of eight molecules of malonyl-CoA to produce octaketides SEK4/SEK4b. As in the case of octaketide synthase, the enzymes are possibly involved in the biosynthesis of the octaketide barbaloin. On the other hand, PKS3 is a multifunctional enzyme that produces a heptaketide aloesone (i.e. the aglycone of aloesin) as a major product from seven molecules of malonyl-CoA. In addition, PKS3 also afforded a hexaketide pyrone (i.e. the precursor of aloenin), a heptaketide 6-(2-acetyl-3,5-dihydroxybenzyl)-4-hydroxy-2-pyrone, a novel heptaketide 6-(2-(2,4-dihydroxy-6-methylphenyl)-2-oxoethyl)-4-hydroxy-2-pyrone and octaketides SEK4/SEK4b. This is the first demonstration of the enzymatic formation of the precursors of the pharmaceutically important aloesin and aloenin by a wild-type PKS obtained from A. arborescens. Interestingly, the aloesone-forming activity was maximum at 50 degrees C, and the novel heptaketide pyrone was non-enzymatically converted to aloesone. In PKS3, the active-site residue 207, which is crucial for controlling the polyketide chain length depending on the steric bulk of the side chain, is uniquely substituted with Ala. Site-directed mutagenesis demonstrated that the A207G mutant dominantly produced the octaketides SEK4/SEK4b, whereas the A207M mutant yielded a pentaketide 5,7-dihydroxy-2-methylchromone.
Subject(s)
Aloe/enzymology , Plant Proteins/chemistry , Polyketide Synthases/chemistry , Aloe/metabolism , DNA, Complementary/chemistry , DNA, Complementary/metabolism , Kinetics , Molecular Sequence Data , Mutagenesis, Site-Directed , Phylogeny , Plant Proteins/classification , Polyketide Synthases/classification , Sequence AlignmentABSTRACT
A C(19) hexaketide stilbene and a C(21) heptaketide chalcone were synthesized by Aloe arborescens octaketide synthase (OKS), a plant-specific type III polyketide synthase (PKS). Remarkably, the C(21) chalcone-forming activity was dramatically increased in a structure-guided OKS N222G mutant that produces a C(20) decaketide SEK15 from 10 molecules of malonyl-CoA. The findings suggested further strategies for production of unnatural polyketides by combination of the precursor-directed biosynthesis and the structure-guided engineering of type III PKS.
Subject(s)
Acyltransferases/metabolism , Aloe/enzymology , Benzophenones/chemical synthesis , Chalcone/chemical synthesis , Stilbenes/chemical synthesis , Aloe/genetics , Benzophenones/chemistry , Catalysis , Chalcone/chemistry , Molecular Structure , Stilbenes/chemistryABSTRACT
A novel aldo-keto reductase (AKR) was cloned and sequenced from roots of Aloe arborescens by a combination of RT-PCR using degenerate primers based on the conserved sequences of plant polyketide reductases (PKRs) and cDNA library screening by oligonucleotide hybridization. A. arborescens AKR share similarities with known plant AKRs (40-66% amino acid sequence identity), maintaining most of the active-site residues conserved in the AKR superfamily enzymes. Interestingly, despite the sequence similarity with PKRs, recombinant enzyme expressed in Escherichia coli did not exhibit any detectable PKR activities. Instead, A. arborescens AKR catalyzed NADPH-dependent reduction of various carbonyl compounds including benzaldehyde and DL-glyceraldehyde. Finally, a homology model on the basis of the crystal structure of Hordeum vulgare AKR predicted the active-site architecture of the enzyme.
Subject(s)
Alcohol Oxidoreductases/genetics , Alcohol Oxidoreductases/metabolism , Aloe/enzymology , Alcohol Oxidoreductases/biosynthesis , Aldehyde Reductase , Aldo-Keto Reductases , Amino Acid Sequence , Cloning, Molecular , DNA Primers , DNA, Complementary/biosynthesis , DNA, Complementary/genetics , DNA, Plant/biosynthesis , DNA, Plant/genetics , Gene Library , Kinetics , Models, Molecular , Molecular Sequence Data , NADP/metabolism , Oligonucleotides , Reverse Transcriptase Polymerase Chain Reaction , Spectrophotometry, UltravioletABSTRACT
Octaketide synthase (OKS) from Aloe arborescens is a plant-specific type III polyketide synthase that produces SEK4 and SEK4b from eight molecules of malonyl-CoA. Recombinant OKS expressed in Escherichia coli was crystallized by the hanging-drop vapour-diffusion method. The crystals belonged to space group I422, with unit-cell parameters a = b = 110.2, c = 281.4 A, alpha = beta = gamma = 90.0 degrees . Diffraction data were collected to 2.6 A resolution using synchrotron radiation at BL24XU of SPring-8.
Subject(s)
Polyketide Synthases/chemistry , Aloe/enzymology , Crystallization , Crystallography, X-RaySubject(s)
Aloe/enzymology , Aloe/metabolism , Drug Design , Macrolides , Polyketide Synthases , Acyltransferases , Amino Acid Sequence , Catalysis , Crystallography, X-Ray , Macrolides/metabolism , Molecular Sequence Data , Polyketide Synthases/chemistry , Polyketide Synthases/physiology , Substrate SpecificityABSTRACT
Pentaketide chromone synthase (PCS) from Aloe arborescens is a novel plant-specific type III polyketide synthase (PKS) that produces 5,7-dihydroxy-2-methylchromone from five molecules of malonyl-CoA. On the basis of the crystal structures of wild-type and M207G mutant PCS, the F80A/Y82A/M207G triple mutant was constructed and shown to produce an unnatural novel nonaketide naphthopyrone by sequential condensations of nine molecules of malonyl-CoA. This is the first demonstration of the formation of a nonaketide by the structurally simple type III PKS. A homology model predicted that the active-site cavity volume of the triple mutant is increased to 4 times that of the wild-type PCS.
Subject(s)
Aloe/enzymology , Polyketide Synthases/chemistry , Protein Engineering , Pyrones/chemistry , Amino Acid Sequence , Base Sequence , Chromatography, High Pressure Liquid , DNA Primers , Molecular Sequence Data , Mutagenesis, Site-Directed , Polyketide Synthases/genetics , Sequence Homology, Amino AcidABSTRACT
The crystal structures of a wild-type and a mutant PCS, a novel plant type III polyketide synthase from a medicinal plant, Aloe arborescens, were solved at 1.6 A resolution. The crystal structures revealed that the pentaketide-producing wild-type and the octaketide-producing M207G mutant shared almost the same overall folding, and that the large-to-small substitution dramatically increases the volume of the polyketide-elongation tunnel by opening a gate to two hidden pockets behind the active site of the enzyme. The chemically inert active site residue 207 thus controls the number of condensations of malonyl-CoA, solely depending on the steric bulk of the side chain. These findings not only provided insight into the polyketide formation reaction, but they also suggested strategies for the engineered biosynthesis of polyketides.
Subject(s)
Aloe/enzymology , Polyketide Synthases/chemistry , Aloe/genetics , Amino Acid Sequence , Amino Acid Substitution , Binding Sites , Chromones , Crystallography, X-Ray , Dimerization , Models, Molecular , Molecular Sequence Data , Mutation , Polyketide Synthases/genetics , Polyketide Synthases/metabolism , Protein Conformation , Protein Structure, Secondary , Sequence AlignmentABSTRACT
Pentaketide chromone synthase (PCS) from Aloe arborescens is a novel plant-specific type III polyketide synthase that catalyzes the formation of 5,7-dihydroxy-2-methylchromone from five molecules of malonyl-CoA. Recombinant PCS expressed in Escherichia coli was crystallized by the hanging-drop vapour-diffusion method. The crystals belonged to space group P2(1), with unit-cell parameters a = 73.2, b = 88.4, c = 70.0 A, alpha = gamma = 90.0, beta = 95.6 degrees . Diffraction data were collected to 1.6 A resolution using synchrotron radiation at BL24XU of SPring-8.
Subject(s)
Aloe/enzymology , Chromones/metabolism , Plant Proteins/chemistry , Polyketide Synthases/chemistry , Chromones/chemistry , Crystallization , Crystallography, X-Ray/methods , Plant Proteins/classification , Plant Proteins/genetics , Polyketide Synthases/classification , Polyketide Synthases/geneticsABSTRACT
We investigated the effect of ultrasound on plasma membrane (PM) Ca2+-ATPase activity of Aloe arborescens callus cells in solid culture. The calluses were exposed by a 20 kHz digital sonifier at the powers of 2 and 10 W from the effective exposure times of 2-10 s. PM Ca2+-ATPase activity was almost significantly higher at 2 W both in continuous wave and 10% duty cycle than that of the control (no ultrasound) at effective exposure times of 5 and 10 s. However, its activity decreased at 10 W in continuous wave exposure. It is possible that the PM Ca2+-ATPase configuration or structure may be partly damaged by high-energy ultrasound at 10 W. Our results showed that low-energy ultrasound exposure was a useful physical field to stimulate A. arborescens callus cells to adapt environmental stress through PM Ca2+-ATPase activity increase.
Subject(s)
Aloe/enzymology , Calcium-Transporting ATPases/metabolism , Cell Membrane/enzymology , Ultrasonics , Aloe/cytology , Aloe/growth & development , Plant Leaves/cytology , Plant Leaves/enzymology , Plant Leaves/growth & developmentABSTRACT
The protective actions of components isolated from Aloe arborescens Miller var. natalensis Berger (Kidachi aloe in Japanese) on streptozotocin (Sz)-induced necrosis of B cells in the pancreatic islets of the mouse were investigated to clarify its action mechanism involved in anti-diabetic effects. In this experiment, phenol low molecular weight components of aloin and aloin A that were anti-oxidants and derived from the leaf skin or pulp extract, an aloe carboxypeptidase fraction that is a inhibitor of enhanced vascular permeability and a glycoprotein component that decreases blood glucose were tested with mice precedently administered with Sz which is known as a cytotoxin specific to B cells. The results showed that the treatment group receiving Sz followed by the aloe carboxypeptidase fraction increased the inhibition of dye leakage by 75.8% (p<0.001) in the extract of whole pancreas in comparison to the control group and the aloe carboxypeptidase fraction group also increased the inhibition effect by 68.4% (p<0.001) in the extract of pancreatic islets as compared to the control group. The carboxypeptidase is an aloe-derived protease known to inhibit the acetic acid-related enhancement of intraperitoneal vascular permeability in mice. Further, the elevation of blood glucose in Sz-induced diabetic mice intraperitoneally given the aloe carboxypeptitase fraction was significantly (p<0.01-0.001) restrained at 3, 7 and 14 days after the injection as compared to the control group given solvent only. The results of this experiment suggested that the inhibitory effect on the enhancement of vascular permeability related to the vascular acute inflammatory response at Sz-induced lesions of pancreatic islets was involved in the action mechanism of this enzyme.
Subject(s)
Aloe/enzymology , Capillary Permeability/drug effects , Carboxypeptidases/pharmacology , Diabetes Mellitus, Experimental/drug therapy , Diabetes Mellitus, Experimental/physiopathology , Islets of Langerhans/drug effects , Animals , Blood Glucose , Carboxypeptidases/metabolism , Dose-Response Relationship, Drug , Female , Male , Mice , Mice, Inbred ICR , Plant Extracts/metabolism , Plant Extracts/pharmacology , Plant Leaves/enzymology , Time FactorsABSTRACT
The chalcone synthase (CHS) superfamily of type III polyketide synthases (PKSs) produces a variety of plant secondary metabolites with remarkable structural diversity and biological activities (e.g., chalcones, stilbenes, benzophenones, acrydones, phloroglucinols, resorcinols, pyrones, and chromones). Here we describe an octaketide-producing novel plant-specific type III PKS from aloe (Aloe arborescens) sharing 50-60% amino acid sequence identity with other plant CHS-superfamily enzymes. A recombinant enzyme expressed in Escherichia coli catalyzed seven successive decarboxylative condensations of malonyl-CoA to yield aromatic octaketides SEK4 and SEK4b, the longest polyketides known to be synthesized by the structurally simple type III PKS. Surprisingly, site-directed mutagenesis revealed that a single residue Gly207 (corresponding to the CHS's active site Thr197) determines the polyketide chain length and product specificity. Small-to-large substitutions (G207A, G207T, G207M, G207L, G207F, and G207W) resulted in loss of the octaketide-forming activity and concomitant formation of shorter chain length polyketides (from triketide to heptaketide) including a pentaketide chromone, 2,7-dihydroxy-5-methylchromone, and a hexaketide pyrone, 6-(2,4-dihydroxy-6-methylphenyl)-4-hydroxy-2-pyrone, depending on the size of the side chain. Notably, the functional diversity of the type III PKS was shown to evolve from simple steric modulation of the chemically inert single residue lining the active-site cavity accompanied by conservation of the Cys-His-Asn catalytic triad. This provided novel strategies for the engineered biosynthesis of pharmaceutically important plant polyketides.
Subject(s)
Acyltransferases/metabolism , Aloe/chemistry , Macrolides/chemical synthesis , Plant Proteins/metabolism , Acyltransferases/chemistry , Aloe/enzymology , Amino Acid Sequence , Macrolides/chemistry , Macrolides/metabolism , Molecular Sequence Data , Molecular Structure , Plant Proteins/chemistry , Plant Roots/chemistry , Protein Biosynthesis , Time FactorsABSTRACT
A novel plant-specific type III polyketide synthase (PKS) that catalyzes formation of a pentaketide chromone, 5,7-dihydroxy-2-methylchromone, from five molecules of malonyl-CoA, was cloned and sequenced from aloe (Aloe arborescens). Site-directed mutagenesis revealed that Met207 (corresponding to Thr197 in CHS) determines the polyketide chain length and the product specificity of the enzyme; remarkably, replacement of a single amino acid residue, Met207, with Gly yielded a mutant enzyme that efficiently produces aromatic octaketides, SEK4 and SEK4b, the products of the minimal PKS for actinorhodin (act from Streptomyces coelicolor), from eight molecules of malonyl-CoA. This provided new insights into the catalytic functions and specificities of the CHS-superfamily type III PKS enzymes.
Subject(s)
Acyltransferases/metabolism , Chromones/metabolism , Plant Proteins/metabolism , Acyltransferases/chemistry , Acyltransferases/genetics , Aloe/enzymology , Aloe/genetics , Amino Acid Sequence , Kinetics , Malonyl Coenzyme A/metabolism , Molecular Sequence Data , Mutagenesis, Site-Directed , Plant Proteins/chemistry , Plant Proteins/genetics , Substrate SpecificityABSTRACT
In Aloe arborescens, an obligate CAM plant, Western analysis detected three major isoforms of NADP-malic enzyme (NADP-ME), 72kDa with a pI of 6.0, 65kDa with a pI of 5.6 and 65kDa with a pI of 5.5. Among them, the 65kDa protein with a pI of 5.5 was leaf-specific, and the 65kDa protein with a pI of 5.6 was found only in roots, whereas the 72kDa protein was uniformly detected in both organs. Activity staining indicated enzyme activity of both 65kDa NADP-MEs but little activity of the 72kDa protein. A cDNA clone encoding a leaf-abundant NADP-ME, AME1, was isolated. Deduced amino acid sequence of AME1 showed a high degree of homology to known NADP-MEs, but it was also found that AME1 contained substitutions on five conservative amino acid residues, some of which have been predicted to be important for their enzyme activity. Transgenic rice carrying the aloe AME1 gene efficiently produced an additional 65kDa protein with a pI of 5.5 as an active NADP-ME. These results indicate that AME1 corresponds to the leaf-specific 65kDa NADP-ME, which may be involved in CAM photosynthesis. It was also shown that substitutions of these conservative amino acid residues identified in AME1 still allowed it to give enzyme activity.
Subject(s)
Aloe/genetics , Amino Acids/genetics , Malate Dehydrogenase/genetics , Plants, Medicinal , Aloe/enzymology , Aloe/metabolism , Amino Acid Sequence , Blotting, Northern , Circadian Rhythm , Conserved Sequence , DNA, Complementary/chemistry , DNA, Complementary/genetics , DNA, Complementary/isolation & purification , Gene Expression Regulation, Enzymologic , Gene Expression Regulation, Plant , Genetic Variation , Isoenzymes/genetics , Molecular Sequence Data , Oryza/genetics , Plant Leaves/enzymology , Plant Leaves/genetics , Plants, Genetically Modified , RNA, Plant/genetics , RNA, Plant/metabolism , Sequence Analysis, DNA , Sequence Homology, Amino AcidABSTRACT
A basic peroxidase (EC 1.11.1.7) (pl around 9.0) has been identified in commercial gel of Aloe barbadensis. In vivo, the activity is localised in the vascular system of inner aqueous leaf parenchyma. Some relevant properties of this basic peroxidase of Aloe have been investigated in leaf extract and in commercial gel where it is notably stable. The acid optimum pH (5.0) for activity and the low KM for H2O2 (0.14 mM) suggest that, when topically applied, Aloe peroxidase may scavenge H2O2 in skin surface.
Subject(s)
Aloe/enzymology , Peroxidases/isolation & purification , Plants, Medicinal , Skin/drug effects , Gels , Peroxidases/pharmacologyABSTRACT
NADP-malic enzyme catalyzes the reaction of decarboxylation from malate. In CAM plants, functions of this enzyme diverged to include both photosynthetic and non-photosynthetic roles. A full length cDNA for an NADP-malic enzyme was isolated from an 'obligate' CAM plant aloe (Aloe arborescens). The cDNA contains an ORF encoding 592 amino acid residues, whose sequence is highly homologous to the known plant NADP-malic enzymes. This gene is constitutively expressed in all organs in a low level. The amount of the transcript exhibited no diurnal variation, suggesting that this gene is not involved in photosynthetic functions.