Ablation of Tmcc2 Gene Impairs Erythropoiesis in Mice.

(1) Background: Transcriptomic and proteomic studies provide a wealth of new genes potentially involved in red blood cell (RBC) maturation or implicated in the pathogenesis of anemias, necessitating validation of candidate genes in vivo; (2) Methods: We inactivated one such candidate, transmembrane and coiled-coil domain 2 (Tmcc2) in mice, and analyzed the erythropoietic phenotype by light microscopy, transmission electron microscopy (TEM), and flow cytometry of erythrocytes and erythroid precursors; (3) Results: Tmcc2-/- pups presented pallor and reduced body weight due to the profound neonatal macrocytic anemia with numerous nucleated RBCs (nRBCs) and occasional multinucleated RBCs. Tmcc2-/- nRBCs had cytoplasmic intrusions into the nucleus and double membranes. Significantly fewer erythroid cells were enucleated. Adult knockouts were normocytic, mildly polycythemic, with active extramedullary erythropoiesis in the spleen. Altered relative content of different stage CD71+TER119+ erythroid precursors in the bone marrow indicated a severe defect of erythroid maturation at the polychromatic to orthochromatic transition stage; (4) Conclusions: Tmcc2 is required for normal erythropoiesis in mice. While several phenotypic features resemble congenital dyserythropoietic anemias (CDA) types II, III, and IV, the involvement of TMCC2 in the pathogenesis of CDA in humans remains to be determined.

Local complications of pentazocine abuse: Case report and review.

The skin is the tissue most commonly affected by intravenous drug addiction with pentazocine. The present article attempts to review the adverse effects of injecting drug use along with one case report of cutaneous complications of injection pentazocine abuse underlining the need for early identification, management, and above all prevention. It also provides credence to the fact that pentazocine abuse is common in paramedical staff, and easy accessibility of pentazocine injection can easily lead to serious complications.

Dimensions of hallucinations and delusions in affective and nonaffective illnesses.

The aim of the study was to examine the dimensions of hallucinations and delusions in affective (manic episode, bipolar affective disorder, and depressive episode) and nonaffective disorders (schizophrenia, acute and transient psychotic disorders, and unspecified psychosis). Sixty outpatients divided equally into two groups comprising affective and nonaffective disorders were taken up for evaluation after screening, as per inclusion and exclusion criteria. Scores of 3 or above on delusion and hallucinatory behavior subscales of positive and negative syndrome scale were sufficient to warrant rating on the psychotic symptom rating scales with which auditory hallucination and delusion were assessed on various dimensions. Insight was assessed using the Beck cognitive insight scale (BCIS). There were no significant differences between the two groups on age, sex, marital status, education, and economic status. There were significant differences in total score and emotional characteristic subscale, cognitive interpretation subscale, and physical characteristic subscale of auditory hallucination scales in between the two groups. Correlation between BCIS-total and total auditory hallucinations score was negative (Spearman Rho -0.319; P < 0.05). Hallucinating patients, more in nonaffective group, described a negative impact of hallucinating voices along with emotional consequences on their lives which lead to distress and disruption.

Agonist-dependent signaling by group I metabotropic glutamate receptors is regulated by association with lipid domains.

Group I metabotropic glutamate receptors (mGluRs), mGluR1 and mGluR5, play critical functions in forms of activity-dependent synaptic plasticity and synapse remodeling in physiological and pathological states. Importantly, in animal models of fragile X syndrome, group I mGluR activity is abnormally enhanced, a dysfunction that may partly underlie cognitive deficits in the condition. Lipid rafts are cholesterol- and sphingolipid-enriched membrane domains that are thought to form transient signaling platforms for ligand-activated receptors. Many G protein-coupled receptors, including group I mGluRs, are present in lipid rafts, but the mechanisms underlying recruitment to these membrane domains remain incompletely understood. Here, we show that mGluR1 recruitment to lipid rafts is enhanced by agonist binding and is supported at least in part by an intact cholesterol recognition/interaction amino acid consensus (CRAC) motif in the receptor. Substitutions of critical residues in the motif reduce mGluR1 association with lipid rafts and agonist-induced, mGluR1-dependent activation of extracellular-signal-activated kinase1/2 MAP kinase (ERK-MAPK). We find that alteration of membrane cholesterol content or perturbation of lipid rafts regulates agonist-dependent activation of ERK-MAPK by group I mGluRs, suggesting a potential function for cholesterol as a positive allosteric modulator of receptor function(s). Together, these findings suggest that drugs that alter membrane cholesterol levels or directed to the receptor-cholesterol interface could be employed to modulate abnormal group I mGluR activity in neuropsychiatric conditions, including fragile X syndrome.

Identification of GPCR localization in detergent resistant membranes.

Lipid domains of the plasma membrane were originally described as a cell matrix insoluble in cold -nonionic detergents and enriched in glycosphingolipids. Because of these biochemical properties, these membrane domains were termed detergent-resistant membranes (DRMs) or detergent-insoluble -glycolipid-enriched (DIG) membranes. Membrane rafts and caveolae are two types of lipid domains that share these properties, as well as structural/functional dependence on membrane cholesterol. Membrane rafts and caveolae are believed to act as signaling platforms for ligand-activated receptors, thereby contributing to the regulation of receptor function. Here we describe a simple method to assess the association of GPCRs with detergent resistant membranes in native brain tissue and cultured cells.

Caveolin-1 knockout mice exhibit impaired induction of mGluR-dependent long-term depression at CA3-CA1 synapses.

Group I metabotropic glutamate receptors (mGluR1/5) are important to synaptic circuitry formation during development and to forms of activity-dependent synaptic plasticity. Dysregulation of mGluR1/5 signaling is implicated in some disorders of neurodevelopment, including fragile X syndrome, the most common inherited form of intellectual disabilities and leading cause of autism. Site(s) in the intracellular loops of mGluR1/5 directly bind caveolin-1, an adaptor protein that associates with membrane rafts. Caveolin-1 is the main coat component of caveolae and organizes macromolecular signaling complexes with effector proteins and membrane receptors. We report that long-term depression (LTD) elicited by a single application of the group I mGluR selective agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) was markedly attenuated at Schaffer collateral-CA1 synapses of mice lacking caveolin-1 (Cav1(-/-)), as assessed by field recording. In contrast, multiple applications of DHPG produced LTD comparable to that in WT mice. Passive membrane properties, basal glutamatergic transmission and NMDA receptor (NMDAR)-dependent LTD were unaltered. The remaining LTD was reduced by anisomycin, an inhibitor of protein synthesis, by U0126, an inhibitor of MEK1/2 kinases, and by rapamycin, an inhibitor of mammalian target of rapamycin (mTOR), suggesting mediation by the same mechanisms as in WT. mGluR1/5-dependent activation (phosphorylation) of MEK and extracellular signal-regulated kinase (ERK1/2) was altered in Cav1(-/-) mice; basal phosphorylation was increased, but a single application of DHPG had no further effect, and after DHPG, phosphorylation was similar in WT and Cav1(-/-) mice. Taken together, our findings suggest that caveolin-1 is required for normal coupling of mGluR1/5 to downstream signaling cascades and induction of mGluR-LTD.

Calreticulin transacylase: genesis, mechanism of action and biological applications.

Our earlier investigations have identified a unique enzyme in the endoplasmic reticulum (ER) termed Acetoxy Drug: Protein Transacetylase (TAase) catalyzing the transfer of acetyl group from polyphenolic acetates (PA) to certain receptor proteins (RP). An elegant assay procedure for TAase was developed based on the inhibition of glutathione S-transferase (GST) due to acetylation by a model acetoxycoumarin, 7, 8-Diacetoxy-4-methylcoumarin (DAMC). TAase purified from various mammalian tissue microsomes to homogeneity exhibited a molecular weight (M.wt) of 55kDa. Further, by N-terminal sequencing TAase was identified as Calreticulin (CR), a multifunctional Ca2+-binding protein in ER lumen. The identity of TAase with CR was evidenced by proteomics studies such as immunoreactivity with anti-CR antibody and mass spectrometry. This function of CR was termed Calreticulin transacetylase (CRTAase). CRTAase was also found to mediate the transfer of acetyl group from DAMC to RP such as NADPH Cytochrome c Reductase (CYPR) and Nitric Oxide Synthase (NOS). The autoacetylation of purified human placental CRTAase concomitant with the acetylation of RP by DAMC was observed. CRTAase activity was found to be inhibited by Ca2+. Our investigations on the individual domains (N, P and C) of CR from a nematode Haemonchus contortus revealed that the P-domain alone was found to possess CRTAase activity. Based on the observation that the autoacetylated CR was a stable intermediate in the CRTAase catalyzed protein acetylation by PA, a putative mechanism was proposed. Further, CRTAase was also found capable of transferring propionyl group from a propoxy derivative of polyphenol, 7,8-Dipropoxy-4-methylcoumarin (DPMC) to RP and concomitant autopropionylation of CR was encountered. Hence, CRTAase was assigned the general term Calreticulin Transacylase. Also, CRTAase was found to act upon the biological acyl group donors, acetyl CoA and propionyl CoA. CRTAase mediated modulation of specific functional proteins by way of acylation was exploited to elicit the biological applications of PA.

Regulation of group I metabotropic glutamate receptor trafficking and signaling by the caveolar/lipid raft pathway.

Endocytic trafficking of neurotransmitter receptors is critical to neuronal signaling and activity-dependent synaptic plasticity. Although the importance of clathrin-mediated endocytosis in receptor trafficking in neurons is well established, the contribution of the caveolar/lipid raft pathway has been little explored. Here, we show that caveolin-1, an adaptor protein that associates with lipid rafts and the main coat protein of caveolae, binds to and colocalizes with metabotropic glutamate receptors 1/5 (mGluR1/5). The interaction with caveolin-1 controls the rate of constitutive mGluR1 internalization, thereby regulating expression of the receptor at the cell surface. Consistent with a role for caveolin-1 in mGluR trafficking, we show that mGluR1/5 associate with lipid rafts in the brain and that their constitutive internalization is mediated, in both heterologous cells and neurons, by caveolar/raft-dependent endocytosis. We further show that caveolin-1 attenuates mGluR1-dependent activation of extracellular signal-regulated kinase (ERK)-mitogen-activated protein kinase (MAPK) signaling, an effect that is abolished in cells expressing mutant mGluR1 lacking intact caveolin binding motifs. Neurons from caveolin-1 knock-out mice show enhanced basal ERK1/2 phosphorylation and prolonged ERK1/2 activation in response to stimulation with DHPG [(RS)-3,5-dihydroxyphenylglycine], a group I mGluR-selective agonist. Together, these findings underscore the importance of caveolar rafts in neurons and suggest that this pathway might play an important role in synapse formation and plasticity.

Proteomic analysis reveals novel binding partners of metabotropic glutamate receptor 1.

Regulated trafficking of neurotransmitter receptors is critical to normal neurodevelopment and neuronal signaling. Group I mGluRs (mGluR1/5 and their splice variants) are G protein-coupled receptors enriched at excitatory synapses, where they serve to modulate glutamatergic transmission. The mGluR1 splice variants mGluR1a and mGluR1b are broadly expressed in the central nervous system and differ in their signaling and trafficking properties. Several proteins have been identified that selectively interact with mGluR1a and participate in receptor trafficking but no proteins interacting with mGluR1b have thus far been reported. We have used a proteomic strategy to isolate and identify proteins that co-purify with mGluR1b in Madin-Darby Canine Kidney (MDCK) cells, an established model system for trafficking studies. Here, we report the identification of 10 novel candidate mGluR1b-interacting proteins. Several of the identified proteins are structural components of the cell cytoskeleton, while others serve as cytoskeleton-associated adaptors and motors or endoplasmic reticulum-associated chaperones. Findings from this work will help unravel the complex cellular mechanisms underlying mGluR trafficking under physiological and pathological conditions.

Autoacetylation of purified calreticulin transacetylase utilizing acetoxycoumarin as the acetyl group donor.

Our earlier reports documented that calreticulin, a multifunctional Ca2+-binding protein in endoplasmic reticulum lumen, possessed protein acetyltransferase function termed Calreticulin Transacetylase (CRTAase). The autoacetylation of purified human placental CRTAase concomitant with the acetylation of receptor proteins by a model acetoxycoumarin, 7,8-Diacetoxy-4-methylcoumarin, was observed. Here, we have examined the autoacetylation property of CRTAase by immunoblotting and mass spectrometry. Ca2+ was found to inhibit CRTAase activity. The inhibition of both autoacetylation of CRTAase as well as acetylation of the receptor protein was apparent when Ca2+) was included in the reaction mixture as visualized by interaction with anti-acetyl lysine antibody. The acetylation of lysines residues: -48, -62, -64, -153, and -159 in N-domain and -206, -207, -209, and -238 in P-domain of CRTAase were located by high-performance liquid chromatography-electronspray ionization tandem mass spectrometry. Further, computer assisted protein structure modeling studies were undertaken to probe the effect of autoacetylation of CRTAase. Accordingly, the predicted CRTAase 3D model showed that all the loop regions of both N- and P-domain bear the acetylated lysines. Energy minimization of the acetylated residues revealed charge neutralization of lysines due to the N-epsilon-acetylation which may facilitate the interaction of CRTAase with the protein substrate and the subsequent transacetylase action.

Establishment of glutamine synthetase of Mycobacterium smegmatis as a protein acetyltransferase utilizing polyphenolic acetates as the acetyl group donors.

Acetoxy Drug: Protein Transacetylase (TAase) mediating the transfer of acetyl group(s) from polyphenolic acetates (PA) to certain functional proteins in mammalian cells was identified by our earlier investigations. TAase activity was characterized in the cell lysates of Mycobacterium smegmatis and the purified protein was found to have M(r) 58,000. TAase catalysed protein acetylation by a model acetoxy drug 7,8-diacetoxy-4-methylcoumarin (DAMC) was established by the demonstration of immunoreactivity of the acetylated target protein with an anti-acetyllysine antibody. The specificity of the TAase of M. smegmatis (MTAase) to various acetoxycoumarins was found to be in the order DAMC > 7-AMC > 6-AMC > 4-AC > 3-AC > ABP. Also, the N-terminal sequence of purified MTAase was found to perfectly match with glutamine synthetase (GS) of M. smegmatis. The identity of MTAase with GS was confirmed by the observation that the purified MTAase as well as the purified recombinant GS exhibited all the properties of GS. The finding that purified Escherichia coli GS was found to have substantial TAase activity highlighted the TAase function of GS in other bacteria. These results conclusively established for the first time the protein acetyltransferase function of GS of M. smegmatis.

Characterization of protein transacetylase from human placenta as a signaling molecule calreticulin using polyphenolic peracetates as the acetyl group donors.

We have earlier shown that a unique membrane-bound enzyme mediates the transfer of acetyl group(s) from polyphenolic peracetates (PA) to functional proteins, which was termed acetoxy drug: protein transacetylase (TAase) because it acted upon several classes of PA. Here, we report the purification of TAase from human placental microsomes to homogeneity with molecular mass of 60 kDa, exhibiting varying degrees of specificity to several classes of PA confirming the structure-activity relationship for the microsome-bound TAase. The TAase catalyzed protein acetylation by a model acetoxy drug, 7,8-diacetoxy-4-methyl coumarin (DAMC) was established by the demonstration of immunoreactivity of the acetylated target protein with anti-acetyl lysine antibody. TAase activity was severely inhibited in calcium-aggregated microsomes as well as when Ca2+ was added to purified TAase, suggesting that TAase could be a calcium binding protein. Furthermore, the N-terminal sequence analysis of purified TAase (EPAVYFKEQFLD) using Swiss Prot Database perfectly matched with calreticulin (CRT), a major microsomal calcium binding protein of the endoplasmic reticulum (ER). The identity of TAase with CRT was substantiated by the observation that the purified TAase avidly reacted with commercially available antibody raised against the C-terminus of human CRT (13 residues peptide, DEEDATGQAKDEL). Purified TAase also showed Ca2+ binding and acted as a substrate for phosphorylation catalyzed by protein kinase C (PKC), which are hallmark characteristics of CRT. Further, purified placental CRT as well as the commercially procured pure CRT yielded significant TAase catalytic activity and were also found effective in mediating the acetylation of the target protein NADPH cytochrome P-450 reductase by DAMC as detected by Western blot using anti-acetyl lysine antibody. These observations for the first time convincingly attribute the transacetylase function to CRT. Hence, this transacetylase function of CRT is designated calreticulin transacetylase (CRTAase). We envisage that CRTAase plays an important role in protein modification by way of acetylation independent of Acetyl CoA.

Acetoxy drug: protein transacetylase catalyzed activation of human platelet nitric oxide synthase by polyphenolic peracetates.

An enhanced intracellular level of Nitric oxide (NO) is essential to ameliorate several pathological conditions of heart and vasculature necessitating the activation of NOS. We have projected in this report the acetylation of eNOS by polyphenolic peracetates (PA) catalyzed by the novel enzyme acetoxy drug: protein transacetylase (TAase) discovered in our laboratory as an unambiguous way of activating NOS which results in the manifestation of physiological action. The human platelet was chosen as the experimental system in order to validate the aforementioned proposition. PA caused profound irreversible activation of platelet NADPH cytochrome c reductase mediated by TAase. The convincing biochemical evidences are presented to show that PA could cause acetylation of the reductase domain of NOS leading to the activation of eNOS in tune with their specificities to platelet TAase. As a result, the enhanced level of NO due to activation of platelet eNOS by PA was found to inhibit the ADP-induced platelet aggregation. The present studies highlight for the first time the role of PA as the novel potent agent for enhancing the intracellular NO levels.

Synthesis of novel amino and acetyl amino-4-methylcoumarins and evaluation of their antioxidant activity.

The six novel 4-methylcoumarins bearing different functionalities such as amino, hydroxy, N-acetyl, acetoxy and nitro have been synthesized and confirmed on the basis of their spectral data (1H-, 13C-NMR, UV, IR and EI mass). They were examined for the first time for their effect on NADPH dependent liver microsomal lipid peroxidation in vitro, and the results were compared with other model 4-methylcoumarin derivatives to establish the structure-activity relationship. Our studies demonstrated that amino group is an effective substitute for the hydroxyl group for antioxidant property and produced a dramatic inhibition of lipid peroxidation. Ortho dihydroxy and ortho hydroxy-amino coumarins were found to possess highest antioxidant and radical scavenging activities.

Acetoxy drug: protein transacetylase of buffalo liver-characterization and mass spectrometry of the acetylated protein product.

The purification and characterization of the buffalo liver microsomal transacetylase (TAase) catalyzing the transfer of acetyl groups from a model acetoxy drug: 7,8-diacetoxy-4-methylcoumarin (DAMC) to GST3-3 has been described here. The enzyme was routinely assayed using DAMC and cytosolic GST as the substrates and was partially purified from microsomes of the buffalo liver. The enzyme was found to have approximate molecular of weight 65 kDa. The action of TAase and DAMC on liver cytosolic GST resulted in the formation of monoacetoxymonohydroxy-4-methylcoumarin (MAMHC) and 7,8-dihydroxy-4-methylcoumarin (DHMC), although the former was the major metabolite. The buffalo liver microsomal TAase exhibited hyperbolic kinetics and yielded K(m) (1667 microM) and V(max) (192 units) when the concentration of DAMC was varied keeping the concentration of GST constant. After having characterized the nature of the substrates and a product of the TAase-catalyzed reaction, we set out to identify the acetylated protein which is another product of the reaction. GST3-3 was used as a model protein substrate for the action of TAase using DAMC as the acetyl donor. The subunit of control and modified GST3-3 were separated by SDS-polyacrylamide gel electrophoresis (PAGE) and digested with trypsin. The tryptic peptides were extracted from the gel pieces and analyzed by matrix assisted laser desorption/ionization-time of flight-mass spectrometry (MALDI-TOFMS). The data search for calibrated and labeled mass peaks of peptides was performed on the Matrix Science Server using the search engine Mascot. The peptide maps so obtained covered 97% of the GST3-3 sequence. On comparison of MALDI peptide maps of modified and control GST, seven new peaks were recognized corresponding to the potentially acetylated peptides in peptide map. The mass value of each of them was 42 Da higher than the theoretical mass of a non-modified GST3-3 tryptic peptide, strongly suggesting acetylation. By examining the fragmentation patterns and by comparing experimental and predicted values for MS/MS daughter ions, the identity of the seven acetylated GST tryptic peptides could be confirmed by the application of LC/MS/MS. In the modified GST, N-terminal proline and six lysines (Lys(51), Lys(82), Lys(123), Lsy(181), Lys(191) and Lys(210)) were found to be acetylated. The structure of acetylated GST revealed that the lysines that underwent acetylation were peripheral in positions.

Mechanism of biochemical action of substituted 4-methylbenzopyran-2-ones. Part 10: identification of inhibitors for the liver microsomal acetoxycoumarin: protein transacetylase.

The quantitative structure-activity relationship (QSAR) studies conducted by us earlier revealed the cardinal role of the pyran ring carbonyl group in the acetoxy polyphenolic compounds for the acetoxy polyphenol:protein transacetylase (TAase) activity. Hence, an attempt was made to examine whether such substrate analogues of benzopyran acetates which lack in the pyran ring carbonyl group, such as 7-acetoxy-2,3-dihydro-2,2-dimethylbenzopyran (BPA), cetachin pentaacetate (CPA) and hematoxylin pentaacetate (HPA) could inhibit the 7,8-diacetoxy-4-methylcoumarin (DAMC):protein (glutathione-S-transferase) transacetylase activity. These compounds were indeed found to remarkably inhibit the TAase activity in a concentration dependent manner and exerted their inhibitory action very rapidly. Further BPA, CPA and HPA were found to abolish the TAase mediated activation of NADPH cytochrome C reductase as well as the inhibition of liver microsome catalyzed aflatoxin B(1) (AFB(1))-DNA binding by DAMC very effectively. These results strongly suggest that the acetoxybenzopyrans merit as potent inhibitors of TAase.

Mechanism of biochemical action of substituted 4-methylbenzopyran-2-ones. Part 9: comparison of acetoxy 4-methylcoumarins and other polyphenolic acetates reveal the specificity to acetoxy drug: protein transacetylase for pyran carbonyl group in proximity to the oxygen heteroatom.

The evidences for the possible enzymatic transfer of acetyl groups (catalyzed by a transacetylase localized in microsomes) from an acetylated compound (acetoxy-4-methylcoumarins) to enzyme proteins leading to profound modulation of their catalytic activities was cited in our earlier publications in this series. The investigations on the specificity for transacetylase (TA) with respect to the number and positions of acetoxy groups on the benzenoid ring of coumarin molecule revealed that acetoxy groups in proximity to the oxygen heteroatom (at C-7 and C-8 positions) demonstrate a high degree of specificity to TA. These studies were extended to the action of TA on acetates of other polyphenols, such as flavonoids and catechin with a view to establish the importance of pyran carbonyl group for the catalytic activity. The absolute requirement of the carbonyl group in the pyran ring of the substrate for TA to function was established by the observation that TA activity was hardly discernible when catechin pentacetate and 7-acetoxy-3,4-dihydro-2,2-dimethylbenzopyran (both lacking pyran ring carbonyl group) were used as the substrates. Further, the TA activity with flavonoid acetates was remarkably lower than that with acetoxycoumarins, thus suggesting the specificity for pyran carbonyl group in proximity to the oxygen heteroatom. The biochemical properties of flavonoid acetates, such as irreversible activation of NADPH cytochrome C reductase and microsome-catalyzed aflatoxin B(1) binding to DNA in vitro were found to be in tune with their specificity to TA.

Establishment of the enzymatic protein acetylation independent of acetyl CoA: recombinant glutathione S-transferase 3-3 is acetylated by a novel membrane-bound transacetylase using 7,8-diacetoxy-4-methyl coumarin as the acetyl donor.

The current knowledge on biological protein acetylation is confined to acetyl CoA-dependent acetylation of protein catalyzed by specific acetyl transferases and the non-enzymatic acetylation of protein by acetylated xenobiotics such as aspirin. We have discovered a membrane-bound enzyme catalyzing the transfer of acetyl groups from the acetyl donor 7,8-diacetoxy-4-methyl coumarin (DAMC) to glutathione S-transferase 3-3 (GST3-3), termed DAMC:protein transacetylase (TAase). The purified enzyme was incubated with recombinant GST3-3 subunit and DAMC, the modified protein was isolated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in gel digested with trypsin and the tryptic digest was analyzed by mass spectrometry. The N-terminus and six lysines, Lys-51, -82, -124, -181, -191 and -210, were found to be acetylated. The acetylation of GST3-3 described above was not observed in the absence of either DAMC or TAase. These results clearly establish the phenomenon of protein acetylation independent of acetyl CoA catalyzed by a hitherto unknown enzyme (TAase) utilizing a certain xenobiotic acetate (DAMC) as the active acetyl donor.

Comparison of the prevention of aflatoxin b(1)-induced genotoxicity by quercetin and quercetin pentaacetate.

Earlier work carried out in our laboratory highlighted the mode of action of acetoxy 4-methylcoumarins in preventing the genotoxicity of aflatoxin B(1) (AFB(1)). We have in this report extended the observations to quercetin pentaacetate (QPA), which unlike quercetin (Q) has demonstrated time-dependent inhibition of liver microsome catalysed AFB(1) epoxidation as measured by AFB(1) binding to DNA. The action of QPA is similar to that of the acetoxy 4-methylcoumarins in that they are acted upon by microsomal transacetylase leading to modulation of catalytic activities of certain enzymes (such as P-450 enzymes, NADPH cytochrome C reductase and glutathione S-transferase) possibly by way of protein acetylation. In the present work, we have documented the transacetylase-mediated action of QPA in preventing genotoxicity due to AFB(1).