scholarly journals Biosynthesis of anandamide and N-palmitoylethanolamine by sequential actions of phospholipase A2 and lysophospholipase D

2004 ◽  
Vol 380 (3) ◽  
pp. 749-756 ◽  
Author(s):  
Yong-Xin SUN ◽  
Kazuhito TSUBOI ◽  
Yasuo OKAMOTO ◽  
Takeharu TONAI ◽  
Makoto MURAKAMI ◽  
...  
Keyword(s):  
Biosynthetic Pathway ◽  
Brain Homogenate ◽  
Wide Distribution ◽  
Rat Tissues ◽  
Lysophospholipase D ◽  
Pla2 Enzyme ◽  
First Time ◽  
Hydrolysis Of ◽  
The Brain

Anandamide (an endocannabinoid) and other bioactive long-chain NAEs (N-acylethanolamines) are formed by direct release from N-acyl-PE (N-acyl-phosphatidylethanolamine) by a PLD (phospholipase D). However, the possible presence of a two-step pathway from N-acyl-PE has also been suggested previously, which comprises (1) the hydrolysis of N-acyl-PE to N-acyl-lysoPE by PLA1/PLA2 enzyme(s) and (2) the release of NAEs from N-acyllysoPE by lysoPLD (lysophospholipase D) enzyme(s). In the present study we report for the first time the characterization of enzymes responsible for this pathway. The PLA1/PLA2 activity for N-palmitoyl-PE was found in various rat tissues, with the highest activity in the stomach. This stomach enzyme was identified as group IB sPLA2 (secretory PLA2), and its product was determined as N-acyl-1-acyl-lysoPE. Recombinant group IB, IIA and V of sPLA2s were also active with N-palmitoyl-PE, whereas group X sPLA2 and cytosolic PLA2α were inactive. In addition, we found wide distribution of lysoPLD activity generating N-palmitoylethanolamine from N-palmitoyl-lysoPE in rat tissues, with higher activities in the brain and testis. Based on several lines of enzymological evidence, the lysoPLD enzyme could be distinct from the known N-acyl-PE-hydrolysing PLD. sPLA2-IB dose dependently enhanced the production of N-palmitoylethanolamine from N-palmitoyl-PE in the brain homogenate showing the lysoPLD activity. N-Arachidonoyl-PE and N-arachidonoyl-lysoPE as anandamide precursors were also good substrates of sPLA2-IB and the lysoPLD respectively. These results suggest that the sequential actions of PLA2 and lysoPLD may constitute another biosynthetic pathway for NAEs, including anandamide.

scholarly journals Assay of advanced glycation endproducts (AGEs): surveying AGEs by chromatographic assay with derivatization by 6-aminoquinolyl-N-hydroxysuccinimidyl-carbamate and application to N∊-carboxymethyl-lysine- and N∊-(1-carboxyethyl)lysine-modified albumin

2002 ◽  
Vol 364 (1) ◽  
pp. 1-14 ◽  
Author(s):  
Naila AHMED ◽  
Ognian K. ARGIROV ◽  
Harjit S. MINHAS ◽  
Carlos A.A. CORDEIRO ◽  
Paul J. THORNALLEY
Keyword(s):  
Advanced Glycation Endproducts ◽  
Advanced Glycation ◽  
Structural Isomers ◽  
Glycation Endproducts ◽  
Assay Procedure ◽  
Carboxymethyl Lysine ◽  
Glycation Process ◽  
First Time ◽  
Hydrolysis Of

Glycation of proteins leads to the formation of early glycation adducts (fructosamine derivatives) and advanced glycation endproducts (AGEs). Formation of AGEs has been linked to the development of cataract, diabetic complications, uraemia, Alzheimer's disease and other disorders. AGEs are a group of compounds of diverse molecular structure and biological function. To characterize AGE-modified proteins used in studies of structural and functional effects of glycation, an assay was developed that surveys the content of early and advanced glycation adducts in proteins. The assay procedure involved enzymic hydrolysis of protein substrate, derivatization of the hydrolysate with 6-aminoquinolyl-N-hydroxysuccinimidyl carbamate (AQC) and HPLC of the resulting adducts with fluorimetric detection. Structural isomers of methylglyoxal-derived hydroimidazolone, glyoxal-derived hydroimidazolone, 3-deoxyglucosone-derived hydroimidazolone and Nδ-(4-carboxy-4,6-dimethyl-5,6-dihydroxy-1,4,5,6-tetrahydropyrimidin-2-yl)-ornithine (THP) were determined for the first time. AGEs with intrinsic fluorescence (argpyrimidine, pentosidine) were assayed without derivatization. Limits of detection were 2–17pmol and levels of recovery were 50–99%, depending on the analyte. The AQC assay resolved structural and epimeric isomers of methylglyoxal-derived hydroimidazolones and THP. Hydroimidazolones, THP and argpyrimidine were AGEs of short-to-intermediate stability under physiological conditions, with half-lives of 1–2weeks. Their measurement provides further insight into the glycation process. The assay was applied to the characterization of human serum albumin minimally and highly modified by N∊-carboxymethyl-lysine and N∊-(1-carboxyethyl)-lysine.

scholarly journals Subcellular localization and tissue distribution of sialic acid-forming enzymes. N-acetylneuraminate-9-phosphate synthase and N-acetylneuraminate 9-phosphatase

1984 ◽  
Vol 223 (2) ◽  
pp. 323-328 ◽  
Author(s):  
J Van Rinsum ◽  
W Van Dijk ◽  
G J Hooghwinkel ◽  
W Ferwerda
Keyword(s):  
Subcellular Localization ◽  
Biosynthetic Pathway ◽  
Cytosolic Fraction ◽  
Phosphate Esters ◽  
Rat Tissues ◽  
Acetylneuraminic Acid ◽  
Nitrophenyl Phosphate ◽  
Sucrose Medium ◽  
Hydrolysis Of ◽  
Phosphate Synthase

The activities of N-acetylneuraminate 9-phosphate synthase and N-acetylneuraminate 9-phosphatase, the two enzymes involved in the final steps of the biosynthetic pathway of N-acetylneuraminic acid, were measured with the substrates N-acetyl[14C]mannosamine 6-phosphate and N-acetyl[14C]neuraminic acid 9-phosphate respectively. Subcellular localization studies in rat liver indicated that both enzymes are localized in the cytosolic fraction after homogenization in sucrose medium. To test the possibility of misinterpretation due to the hydrolysis of N-acetylneuraminic acid 9-phosphate by non-specific phosphatases, the hydrolysis of various phosphate esters by the cytosolic fraction was tested. Only p-nitrophenyl phosphate was hydrolysed; however, competition studies with N-acetylneuraminic acid 9-phosphate and p-nitrophenyl phosphate indicated that two different enzymes were involved and that no competition existed between the two substrates. In various other rat tissues N-acetylneuraminate-9-phosphate synthase and N-acetylneuraminate 9-phosphatase activities were detected, suggesting that N-acetylmannosamine 6-phosphate is a general precursor for N-acetylneuraminic acid biosynthesis in all the tissues studied.

Fabrication and Characterization of Biodegradable Metal Based Microelectrodes for In Vivo Neural Recording

MRS Advances ◽  
2019 ◽  
Vol 4 (46-47) ◽  
pp. 2471-2477
Author(s):  
Chaoxing Zhang ◽  
Teresa H. Wen ◽  
Khaleel A. Razak ◽  
Jiajia Lin ◽  
Edgar Villafana ◽  
...  
Keyword(s):  
Neural Recording ◽  
Neural Signals ◽  
Secondary Surgery ◽  
New Class ◽  
Fabrication And Characterization ◽  
Evoked Activity ◽  
First Time ◽  
The Brain

ABSTRACT:Neural electrodes have been widely used to monitor neural signals and/or deliver electrical stimulation in the brain. Currently, biodegradable and biocompatible materials have been actively investigated to create temporary electrodes that could degrade after serving their functions for neural recording and stimulation from days to months. The new class of biodegradable electrodes eliminate the necessity of secondary surgery for electrode removal. In this study, we created biodegradable, biocompatible, and implantable magnesium (Mg)-based microelectrodes for in vivo neural recording for the first time. Specifically, conductive poly-3,4-ethylenedioxythiophene (PEDOT) was first deposited onto Mg microwire substrates by electrochemical deposition, and a biodegradable insulating polymer was subsequently sprayed onto the surface of electrodes. The tip of electrodes was designed to be conductive for neural recording and stimulation, while the rest of electrodes was insulated with a polymer that is biocompatible with neural tissue. The impedance of Mg-based microelectrodes and their performance during neural recording in the auditory cortex of a mouse were studied. The results first demonstrated the capability of Mg-based microelectrodes for in vivo recording of multi-unit stimulus-evoked activity in the brain.

scholarly journals Characterization of the CDP-2-Glycerol Biosynthetic Pathway in Streptococcus pneumoniae

2010 ◽  
Vol 192 (20) ◽  
pp. 5506-5514 ◽  
Author(s):  
Quan Wang ◽  
Yanli Xu ◽  
Andrei V. Perepelov ◽  
Wei Xiong ◽  
Dongmei Wei ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Streptococcus Pneumoniae ◽  
Biosynthetic Pathway ◽  
Resonance Spectroscopy ◽  
Ionization Mass ◽  
Enzyme Substrate ◽  
Effects Of Temperature ◽  
First Time ◽  
Novel Products

ABSTRACT Capsule polysaccharide (CPS) plays an important role in the virulence of Streptococcus pneumoniae and is usually used as the pneumococcal vaccine target. Glycerol-2-phosphate is found in the CPS of S. pneumoniae types 15A and 23F and is rarely found in the polysaccharides of other bacteria. The biosynthetic pathway of the nucleotide-activated form of glycerol-2-phosphate (NDP-2-glycerol) has never been identified. In this study, three genes (gtp1, gtp2, and gtp3) from S. pneumoniae 23F that have been proposed to be involved in the synthesis of NDP-2-glycerol were cloned and the enzyme products were expressed, purified, and assayed for their respective activities. Capillary electrophoresis was used to detect novel products from the enzyme-substrate reactions, and the structure of the product was elucidated using electrospray ionization mass spectrometry and nuclear magnetic resonance spectroscopy. Gtp1 was identified as a reductase that catalyzes the conversion of 1,3-dihydroxyacetone to glycerol, Gtp3 was identified as a glycerol-2-phosphotransferase that catalyzes the conversion of glycerol to glycerol-2-phosphate, and Gtp2 was identified as a cytidylyltransferase that transfers CTP to glycerol-2-phosphate to form CDP-2-glycerol as the final product. The kinetic parameters of Gtp1 and Gtp2 were characterized in depth, and the effects of temperature, pH, and cations on these two enzymes were analyzed. This is the first time that the biosynthetic pathway of CDP-2-glycerol has been identified biochemically; this pathway provides a method to enzymatically synthesize this compound.

Preparation and Characterization of Novel Brain Targeting Magnetic Nanoparticles Modified with Ascorbic Acid

NANO ◽  
2018 ◽  
Vol 13 (01) ◽  
pp. 1850008 ◽  
Author(s):  
Linhui Wang ◽  
Yi Zhao ◽  
Runxin Lu ◽  
Yao Peng ◽  
Li Guo ◽  
...  
Keyword(s):  
Ascorbic Acid ◽  
Magnetic Nanoparticles ◽  
Glucose Transporter ◽  
Drug Loading ◽  
Brain Homogenate ◽  
Brain Targeting ◽  
Glucose Transporter 1 ◽  
Cns Drug Delivery ◽  
The Brain

Magnetic targeting, which utilizes a magnetic field to specifically deliver therapeutic agents to the targeted regions, can greatly improve the treatment efficiency. Herein, ibuprofen-loaded brain targeting magnetic nanoparticles (AA-Ibu-PEG-DA@MNPs) modified with ascorbic acid (AA) for central nervous system (CNS) drug delivery was designed and synthesized in order to effectively deliver ibuprofen to the brain through Na[Formula: see text]-dependent vitamin C transporter 2 (SVCT2) and glucose transporter 1 (GLUT1). The brain targeting magnetic nanoparticles, AA-Ibu-PEG-DA@MNPs, have a particle size of 82.5[Formula: see text]nm, 2% drug loading capacity and limited cytotoxicity against bEnd.3 cells. What’s more, the nanoparticles maintained the magnetic property with a saturation magnetization level at 52.17[Formula: see text]emu/g and could release ibuprofen when incubated in different mediums, including various buffers, mice plasma and brain homogenate. The results indicate that the magnetic nanoparticles may have the potential to be a promising approach to selectively deliver drugs into the brain. This study may be conducive to the field of CNS drugs delivery.

scholarly journals Cryoenzymology of trypsin. 13C-n.m.r. detection of an acyl-trypsin intermediate in the trypsin-catalysed hydrolysis of a highly specific substrate at subzero temperature

1984 ◽  
Vol 219 (2) ◽  
pp. 437-444 ◽  
Author(s):  
N E Mackenzie ◽  
J P Malthouse ◽  
A I Scott
Keyword(s):  
Subzero Temperature ◽  
Specific Substrate ◽  
Visible Absorption ◽  
Enzyme Substrate ◽  
First Time ◽  
Kinetics Of ◽  
Hydrolysis Of ◽  
Nitrophenyl Ester

The kinetics of the trypsin-catalysed hydrolysis of the highly specific substrate N alpha-benzyloxycarbonyl-L-lysine p-nitrophenyl ester were studied under cryoenzymological conditions by 13C-n.m.r. spectroscopy at pH approx. 3.0. The kinetics of this reaction are shown to be in agreement with similar studies made with the use of u.v.-visible-absorption-spectrophotometric techniques. A combination of 13C-n.m.r. spectroscopy and cryoenzymology has for the first time detected an acyl-trypsin intermediate in the hydrolysis of this highly specific substrate. The advantages and difficulties of using 13C-n.m.r. spectroscopy coupled with cryoenzymology in the detection and characterization of enzyme-substrate intermediates are discussed.

scholarly journals Characterization of Mariner transposons in seven species of Rhus gall aphids

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Aftab Ahmad ◽  
Gabriel Luz Wallau ◽  
Zhumei Ren
Keyword(s):  
Transposable Elements ◽  
Present Report ◽  
Wide Distribution ◽  
Inverted Repeats ◽  
Genomic Evolution ◽  
Reading Frame ◽  
Jumping Genes ◽  
First Time ◽  
Intact Open Reading Frame

AbstractTransposable elements (TEs), also known as jumping genes, are widely spread in the genomes of insects and play a considerable role in genomic evolution. Mariner/DD34D family belongs to class II transposable elements which is widely spread in the genomes of insects and have considerable role in genomic evolution. Mariner like elements (MLEs) were searched in the genomes of seven species of Rhus gall aphids belonging to six genera. In total, 121 MLEs were detected in the genomes of the seven investigated species of Rhus gall aphids, which showed a wide distribution in both close and distant related species. The sequences of MLEs ranged from 1 to 1.4 kb in length and the structural analysis of the MLEs showed that only five copies were potentially active with intact open reading frame (ORF) and terminal inverted repeats (TIRs). Phylogenetic analysis showed that all the 121 MLE sequences belonged to four subfamilies, i.e., Mauritiana, Drosophila, Vertumana and Irritans, among which Drosophila and Vertumana subfamilies were reported in aphids for the first time. Our present report revealed the diversity and distribution of MLEs in Rhus gall aphid genomes and expanded our understandings on the characterization of transposable elements in aphid genomes, which might be useful as genetic markers and tools and would play an important role in genomic evolution and adaptation of aphids.

scholarly journals Enzymatic Conversion of Oleuropein to Hydroxytyrosol Using Immobilized β-Glucosidase on Porous Carbon Cuboids

Nanomaterials ◽  
2019 ◽  
Vol 9 (8) ◽  
pp. 1166 ◽  
Author(s):  
Chatzikonstantinou ◽  
Gkantzou ◽  
Thomou ◽  
Chalmpes ◽  
Lyra ◽  
...  
Keyword(s):  
Porous Carbon ◽  
Photoelectron Spectroscopy ◽  
Thermotoga Maritima ◽  
Covalent Immobilization ◽  
Force Microscopy ◽  
Atomic Force ◽  
Repeated Use ◽  
First Time ◽  
Hydrolysis Of

In the present study, we developed novel β-glucosidase-based nano-biocatalysts for the bioconversion of oleuropein to hydroxytyrosol. Using non-covalent or covalent immobilization approaches, β-glucosidases from almonds and Thermotoga maritima were attached for the first time on oxidized and non-oxidized porous carbon cuboids (PCC). Various methods were used for the characterization of the bio-nanoconjugates, such as Fourier transform infrared spectroscopy (FTIR), X-ray photoelectron spectroscopy (XPS), atomic force microscopy (AFM), and fluorescence spectroscopy. The oxidation state of the nanο-support and the immobilization procedure play a key role for the immobilization efficiency or the catalytic activity of the immobilized β-glucosidases. The nano-biocatalysts were successfully used for the hydrolysis of oleuropein, which leads to the formation of its bioactive derivative, hydroxytyrosol (up to 2.4 g L−1), which is a phenolic compound with numerous health benefits. The bio-nanoconjugates exhibited high thermal and operational stability (up to 240 hours of repeated use), which indicated that they are efficient tools for various bio-transformations.

scholarly journals Characterization of Mariner transposons in Rhus gall aphids (Hemiptera: Aphididae: Eriosomatinae)

2021 ◽  
Author(s):  
Aftab Ahmad ◽  
Gabriel Luz Wallau ◽  
Zhumei Ren
Keyword(s):  
Transposable Elements ◽  
Present Report ◽  
Wide Distribution ◽  
Genomic Evolution ◽  
Reading Frame ◽  
Jumping Genes ◽  
The One ◽  
First Time ◽  
Intact Open Reading Frame

Abstract Background: Transposable elements (TEs), also known as jumping genes, are widely spread in the genomes of insects and play a considerable role in genomic evolution. Mariner family belongs to class II transposable elements, were searched in the genomes of seven species of Rhus gall aphids belonging to six genera. Mariner-like elements were characterized for the first time in Rhus gall aphids and classified in to respective subfamilies.Results: In total, one hundred twenty-one MLEs were detected in the genomes of the seven investigated species of Rhus gall aphids, which showed a wide distribution of MLEs in both close and distant related species. The sequences of MLEs ranged from 1kb to 1.4kb in length and the structural analysis of the MLEs showed that only five copies were potentially active with intact open reading frame (ORF) while the remaining were classified as inactive MLEs according to absence of single intact ORF or terminal inverted repeats (TIRs). Based on the MLEs in Rhus gall aphids as well as the well characterized MLEs in other organisms from GenBank, the phylogenetic analysis showed that all the one hundred twenty-one MLE sequences belonged to four subfamilies, i.e., thirty from Maurutiana subfamily, twenty-six from Drosophila subfamily, thirty-three from Vertumana subfamily and thirty-two from Irritans subfamily, among which Drosophila and Vertumana subfamilies were reported in aphids for the first time. Moreover, the phylogenetic relationship suggested possible horizontal transfer events of MLEs between aphids and other insects.Conclusion: Our present report revealed the diversity and distribution of MLEs in Rhus gall aphid genomes sequenced by shotgun genome skimming method. This study further expanded our understandings on the characterization of transposable elements in aphid genomes, which might be useful as genetic markers and tools and would play an important role in genomic evolution and adaptation of aphids.

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