The Experts below are selected from a list of 3423 Experts worldwide ranked by ideXlab platform
Jose L Daniotti - One of the best experts on this subject based on the ideXlab platform.
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individual s acylated cysteines differentially contribute to h ras endomembrane trafficking and acylation Deacylation cycles
Molecular Biology of the Cell, 2017Co-Authors: Maria P. Pedro, Guillermo A Gomez, Aldo Alejandro Vilcaes, Jose L DaniottiAbstract:S-acylation/Deacylation cycles and vesicular transport are critical for an adequate subcellular distribution of S-acylated Ras proteins. H-Ras is dually acylated on cysteines 181 and 184, but it is unknown how these residues individually contribute to H-Ras traffcking. In this study, we characterized the acylation and Deacylation rates and membrane traffcking of monoacylated H-Ras mutants to analyze their contributions to H-Ras plasma membrane and endomembrane distribution. We demonstrated that dually acylated H-Ras interacts with acylprotein thioesterases (APTs) 1 and 2 at the plasma membrane. Moreover, single-acylation mutants of H-Ras differed not only in their subcellular distribution, where both proteins localized to different extents at both the Golgi complex and plasma membrane, but also in their Deacylation rates, which we showed to be due to different sensitivities to APT1 and APT2. Fluorescence photobleaching and photoactivation experiments also revealed that 1) although S-acylated, single-acylation mutants are incorporated with different effciencies into Golgi complex to plasma membrane vesicular carriers, and 2) the different Deacylation rates of single-acylated H-Ras influence differentially its overall exchange between different compartments by nonvesicular transport. Taken together, our results show that individual S-acylation sites provide singular information about H-Ras subcellular distribution that is required for GTPase signaling.
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2-Bromopalmitate Reduces Protein Deacylation by Inhibition of Acyl-Protein Thioesterase Enzymatic Activities
2016Co-Authors: Maria P. Pedro, Vanesa M Tomatis, Guillermo A Gomez, Aldo A. Vilcaes, Rafael G. Oliveira, Jose L DaniottiAbstract:S-acylation, the covalent attachment of palmitate and other fatty acids on cysteine residues, is a reversible post-translational modification that exerts diverse effects on protein functions. S-acylation is catalyzed by protein acyltransferases (PAT), while Deacylation requires acyl-protein thioesterases (APT), with numerous inhibitors for these enzymes having already been developed and characterized. Among these inhibitors, the palmitate analog 2-brompalmitate (2-BP) is the most commonly used to inhibit palmitoylation in cells. Nevertheless, previous results from our laboratory have suggested that 2-BP could affect protein Deacylation. Here, we further investigated in vivo and in vitro the effect of 2-BP on the acylation/Deacylation protein machinery, with it being observed that 2-BP, in addition to inhibiting PAT activity in vivo, also perturbed the acylation cycle of GAP-43 at the level of depalmitoylation and consequently affected its kinetics of membrane association. Furthermore, 2-BP was able to inhibit in vitro the enzymatic activities of human APT1 and APT2, the only two thioesterases shown to mediate protein Deacylation, through an uncompetitive mechanism of action. In fact, APT1 and APT2 hydrolyzed both the monomeric form as well as the micellar state of the substrate palmitoyl-CoA. On the basis of the obtained results, as APTs can mediate Deacylation on membrane bound and unbound substrates, this suggests that the access of APTs to the membrane interface is not a necessary requisite for Deacylation. Moreover, as the enzymatic activity of APTs was inhibited by 2-BP treatment, then the kinetics analysis of protein acylation using 2-BP should be carefully interpreted, as this drug als
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acyl protein thioesterase 2 catalizes the Deacylation of peripheral membrane associated gap 43
PLOS ONE, 2010Co-Authors: Vanesa M Tomatis, Alejandra Trenchi, Guillermo A Gomez, Jose L DaniottiAbstract:An acylation/Deacylation cycle is necessary to maintain the steady-state subcellular distribution and biological activity of S-acylated peripheral proteins. Despite the progress that has been made in identifying and characterizing palmitoyltransferases (PATs), much less is known about the thioesterases involved in protein Deacylation. In this work, we investigated the Deacylation of growth-associated protein-43 (GAP-43), a dually acylated protein at cysteine residues 3 and 4. Using fluorescent fusion constructs, we measured in vivo the rate of Deacylation of GAP-43 and its single acylated mutants in Chinese hamster ovary (CHO)-K1 and human HeLa cells. Biochemical and live cell imaging experiments demonstrated that single acylated mutants were completely deacylated with similar kinetic in both cell types. By RT-PCR we observed that acyl-protein thioesterase 1 (APT-1), the only bona fide thioesterase shown to mediate Deacylation in vivo, is expressed in HeLa cells, but not in CHO-K1 cells. However, APT-1 overexpression neither increased the Deacylation rate of single acylated GAP-43 nor affected the steady-state subcellular distribution of dually acylated GAP-43 both in CHO-K1 and HeLa cells, indicating that GAP-43 Deacylation is not mediated by APT-1. Accordingly, we performed a bioinformatic search to identify putative candidates with acyl-protein thioesterase activity. Among several candidates, we found that APT-2 is expressed both in CHO-K1 and HeLa cells and its overexpression increased the Deacylation rate of single acylated GAP-43 and affected the steady-state localization of diacylated GAP-43 and H-Ras. Thus, the results demonstrate that APT-2 is the protein thioesterase involved in the acylation/Deacylation cycle operating in GAP-43 subcellular distribution.
Kevin J. Edgar - One of the best experts on this subject based on the ideXlab platform.
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Probing the Mechanism of TBAF-Catalyzed Deacylation of Cellulose Esters
2016Co-Authors: Xueyan Zheng, Richard D. Gandour, Kevin J. EdgarAbstract:The mechanism of the recently discovered, unusual tetrabutylammonium fluoride (TBAF) Deacylation of cellulose esters has been investigated by methods including kinetic isotope effect (KIE) studies. The secondary KIE (kH/kD = 1.26 ± 0.04) measured for Deacylation at C-2/3 suggests a mechanism involving a ketene intermediate for those positions. An inverse KIE (kH/kD = 0.87 ± 0.03) for the Deacylation at C-6 indicates the involvement of a tetrahedral intermediate in the mechanism. Additional studies suggest the possibility that TBAF chelation by neighboring acyl groups may account for the unexpected regioselectivity at the secondary alcohol esters that is observed in the TBAF Deacylation of cellulose esters
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TBAF and Cellulose Esters: Unexpected Deacylation with Unexpected Regioselectivity
2016Co-Authors: Kevin J. EdgarAbstract:Tetrabutylammonium fluoride has been found to catalyze the Deacylation of cellulose esters. More surprisingly, the Deacylation is highly regioselective. Even more remarkably, in contrast with the C-6 regioselectivity of other reactions of cellulose and its derivatives, this Deacylation shows substantial selectivity for the removal of the acyl groups from the esters of the secondary alcohols at C-2 and C-3, affording cellulose-6-O-esters with high regioselectivity by a simple one-step process employing no protective groups
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remarkably regioselective Deacylation of cellulose esters using tetraalkylammonium salts of the strongly basic hydroxide ion
Carbohydrate Polymers, 2014Co-Authors: Xueyan Zheng, Richard D. Gandour, Kevin J. EdgarAbstract:Tetraalkylammonium hydroxides have been found to mediate regioselective Deacylation of cellulose esters. This Deacylation surprisingly shows substantial selectivity for the removal of the acyl groups at O-2/3, affording cellulose-6-O-esters by a simple, efficient one-step process. The mechanism for this Deacylation was investigated by studying the effect of tetraalkylammonium cation size upon ester Deacylation selectivity. We hypothesize that coordination of the tetraalkylammonium cation by the ester oxygen atoms of the vicinal 2,3-acetate groups may drive the unexpected regioselectivity at the secondary alcohol esters. Broad scope with respect to ester type was demonstrated; regioselective O-2,3 Deacylation was observed with cellulose acetate, propionate, butyrate, hexanoate and benzoate triesters. The scope of this Deacylation of cellulose acetates has been investigated to understand how to carry it out most efficiently. Reaction with TBAOH in pyridine was the most effective process, providing the highest selectivity.
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tbaf catalyzed Deacylation of cellulose esters reaction scope and influence of reaction parameters
Carbohydrate Polymers, 2013Co-Authors: Xueyan Zheng, Richard D. Gandour, Kevin J. EdgarAbstract:Abstract In order to expand its utility and understand how to carry it out most efficiently, the scope of the highly regioselective, tetrabutylammonium fluoride (TBAF) catalyzed Deacylation of cellulose acetates has been investigated, including the influence of key process parameters: solvent, temperature, and water content. Reactions in DMSO, THF, MEK and acetone afforded similar extents of Deacylation and regioselectivity. Reaction with TBAF in DMSO at 50 °C for 18 h was the most efficient process providing regioselective Deacylation at O-2/3. All results were consistent with our previous mechanistic proposals. Furthermore, we demonstrate that TBAF-catalyzed Deacylation is also effective and regioselective with cellulose acetate, butyrate, and hexanoate triesters, and even with a cellulose ester devoid of alpha protons, cellulose tribenzoate. These reactions displayed regioselectivity for Deacylation at O-2/3 similar to that observed earlier with cellulose acetate (DS 2.4).
Kazushi Mashima - One of the best experts on this subject based on the ideXlab platform.
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a tetranuclear zinc cluster catalyzed practical and versatile deprotection of acetates and benzoates
ChemInform, 2011Co-Authors: Takanori Iwasaki, Kazushi Agura, Yusuke Maegawa, Yukiko Hayashi, Takashi Ohshima, Kazushi MashimaAbstract:A new catalytic Deacylation of acetates and benzoates through transesterification with methanol is developed.
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a tetranuclear zinc cluster catalyzed practical and versatile deprotection of acetates and benzoates
Chemistry: A European Journal, 2010Co-Authors: Takanori Iwasaki, Kazushi Agura, Yusuke Maegawa, Yukiko Hayashi, Takashi Ohshima, Kazushi MashimaAbstract:A new catalytic Deacylation of acetates and benzoates through transesterification with methanol was developed (see scheme). Reactions with various acid- and nucleophile-sensitive functional groups proceeded efficiently in the presence of a catalytic amount of the tetranuclear zinc cluster. The present catalysis is applicable to less-reactive tertiary acetates, the Deacylation of which is difficult to achieve by transesterification with other catalysts.
Yi Qin Gao - One of the best experts on this subject based on the ideXlab platform.
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a click chemistry approach reveals the chromatin dependent histone h3k36 deacylase nature of sirt7
Journal of the American Chemical Society, 2019Co-Authors: Wesley Wei Wang, Jie Lyu, Yadagiri Kurra, Zhen Tong, Ling Zhang, Vangmayee Sharma, Jennifer Zhou, Hening Lin, Maria Anguloibanez, Yi Qin GaoAbstract:Using an engineered pyrrolysyl-tRNA synthetase mutant together with tRNACUAPyl, we have genetically encoded Ne-(7-azidoheptanoyl)-l-lysine (AzHeK) by amber codon in Escherichia coli for recombinant expression of a number of AzHeK-containing histone H3 proteins. We assembled in vitro acyl-nucleosomes from these recombinant acyl-H3 histones. All these acyl-nucleosomes contained an azide functionality that allowed quick click labeling with a strained alkyne dye for in-gel fluorescence analysis. Using these acyl-nucleosomes as substrates and click labeling as a detection method, we systematically investigated chromatin Deacylation activities of SIRT7, a class III NAD+-dependent histone deacylase with roles in aging and cancer biology. Besides confirming the previously reported histone H3K18 Deacylation activity, our results revealed that SIRT7 has an astonishingly high activity to catalyze Deacylation of H3K36 and is also catalytically active to deacylate H3K37. We further demonstrated that this H3K36 Deacylation activity is nucleosome dependent and can be significantly enhanced when appending the acyl-nucleosome substrate with a short double-stranded DNA that mimics the bridging DNA between nucleosomes in native chromatin. By overexpressing SIRT7 in human cells, we verified that SIRT7 natively removes acetylation from histone H3K36. Moreover, SIRT7-deficient cells exhibited H3K36 hyperacetylation in whole cell extracts, at rDNA sequences in nucleoli, and at select SIRT7 target loci, demonstrating the physiologic importance of SIRT7 in determining endogenous H3K36 acetylation levels. H3K36 acetylation has been detected at active gene promoters, but little is understood about its regulation and functions. Our findings establish H3K36 as a physiologic substrate of SIRT7 and implicate this modification in potential SIRT7 pathways in heterochromatin silencing and genomic stability.
Xueyan Zheng - One of the best experts on this subject based on the ideXlab platform.
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Probing the Mechanism of TBAF-Catalyzed Deacylation of Cellulose Esters
2016Co-Authors: Xueyan Zheng, Richard D. Gandour, Kevin J. EdgarAbstract:The mechanism of the recently discovered, unusual tetrabutylammonium fluoride (TBAF) Deacylation of cellulose esters has been investigated by methods including kinetic isotope effect (KIE) studies. The secondary KIE (kH/kD = 1.26 ± 0.04) measured for Deacylation at C-2/3 suggests a mechanism involving a ketene intermediate for those positions. An inverse KIE (kH/kD = 0.87 ± 0.03) for the Deacylation at C-6 indicates the involvement of a tetrahedral intermediate in the mechanism. Additional studies suggest the possibility that TBAF chelation by neighboring acyl groups may account for the unexpected regioselectivity at the secondary alcohol esters that is observed in the TBAF Deacylation of cellulose esters
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remarkably regioselective Deacylation of cellulose esters using tetraalkylammonium salts of the strongly basic hydroxide ion
Carbohydrate Polymers, 2014Co-Authors: Xueyan Zheng, Richard D. Gandour, Kevin J. EdgarAbstract:Tetraalkylammonium hydroxides have been found to mediate regioselective Deacylation of cellulose esters. This Deacylation surprisingly shows substantial selectivity for the removal of the acyl groups at O-2/3, affording cellulose-6-O-esters by a simple, efficient one-step process. The mechanism for this Deacylation was investigated by studying the effect of tetraalkylammonium cation size upon ester Deacylation selectivity. We hypothesize that coordination of the tetraalkylammonium cation by the ester oxygen atoms of the vicinal 2,3-acetate groups may drive the unexpected regioselectivity at the secondary alcohol esters. Broad scope with respect to ester type was demonstrated; regioselective O-2,3 Deacylation was observed with cellulose acetate, propionate, butyrate, hexanoate and benzoate triesters. The scope of this Deacylation of cellulose acetates has been investigated to understand how to carry it out most efficiently. Reaction with TBAOH in pyridine was the most effective process, providing the highest selectivity.
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tbaf catalyzed Deacylation of cellulose esters reaction scope and influence of reaction parameters
Carbohydrate Polymers, 2013Co-Authors: Xueyan Zheng, Richard D. Gandour, Kevin J. EdgarAbstract:Abstract In order to expand its utility and understand how to carry it out most efficiently, the scope of the highly regioselective, tetrabutylammonium fluoride (TBAF) catalyzed Deacylation of cellulose acetates has been investigated, including the influence of key process parameters: solvent, temperature, and water content. Reactions in DMSO, THF, MEK and acetone afforded similar extents of Deacylation and regioselectivity. Reaction with TBAF in DMSO at 50 °C for 18 h was the most efficient process providing regioselective Deacylation at O-2/3. All results were consistent with our previous mechanistic proposals. Furthermore, we demonstrate that TBAF-catalyzed Deacylation is also effective and regioselective with cellulose acetate, butyrate, and hexanoate triesters, and even with a cellulose ester devoid of alpha protons, cellulose tribenzoate. These reactions displayed regioselectivity for Deacylation at O-2/3 similar to that observed earlier with cellulose acetate (DS 2.4).