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Carbohydrate Polymers Dec 2023Galactoglucomannans (GGM) recovered from abundant forest industry side-streams has been widely recognized as a renewable hydrocolloid. The low molar mass and presence of...
Galactoglucomannans (GGM) recovered from abundant forest industry side-streams has been widely recognized as a renewable hydrocolloid. The low molar mass and presence of O-acetyl side-groups results in low viscous dispersions and weak intermolecular interactions that make GGM unsuitable for hydrogel formation, unless forcefully chemically derivatized and/or crosslinked with other polymers. Here we present the characterization of hydrogels prepared from GGM after tailoring the degree of acetylation by alkaline treatment during its recovery. Specifically, we investigated gel characteristics of low-acetyl GGM dispersions prepared at varied solid concentrations (5, 10 and 15 %) and pH (4, 7 and 10), and then subjected to ultrasonication. The results indicated that low-acetyl GGM dispersions formed gels (G' > G″) at all other studied solid concentration and pH level combinations except 5 % and pH 4. High pH levels, leading to further removal of acetyl groups, and high solid concentration facilitated the gel formation. GGM hydrogels were weak gels with strong shear-thinning behavior and thixotropic properties, and high hardness and water holding capacity; which were enhanced with increased pH and solid concentration, and prolonged storage time. Our study showed the possibility to utilize low-acetyl GGM as mildly processed gelling or thickening agents, and renewable materials for bio-based hydrogels.
Topics: Hydrogels; Mannans; Acetylation; Hardness; Picea
PubMed: 37739540
DOI: 10.1016/j.carbpol.2023.121316 -
Communications Biology Sep 2023Cytosolic citrate is imported from the mitochondria by SLC25A1, and from the extracellular milieu by SLC13A5. In the cytosol, citrate is used by ACLY to generate...
Cytosolic citrate is imported from the mitochondria by SLC25A1, and from the extracellular milieu by SLC13A5. In the cytosol, citrate is used by ACLY to generate acetyl-CoA, which can then be exported to the endoplasmic reticulum (ER) by SLC33A1. Here, we report the generation of mice with systemic overexpression (sTg) of SLC25A1 or SLC13A5. Both animals displayed increased cytosolic levels of citrate and acetyl-CoA; however, SLC13A5 sTg mice developed a progeria-like phenotype with premature death, while SLC25A1 sTg mice did not. Analysis of the metabolic profile revealed widespread differences. Furthermore, SLC13A5 sTg mice displayed increased engagement of the ER acetylation machinery through SLC33A1, while SLC25A1 sTg mice did not. In conclusion, our findings point to different biological responses to SLC13A5- or SLC25A1-mediated import of citrate and suggest that the directionality of the citrate/acetyl-CoA pathway can transduce different signals.
Topics: Animals; Mice; Acetyl Coenzyme A; Acetylation; Citrates; Citric Acid; Phenotype
PubMed: 37689798
DOI: 10.1038/s42003-023-05311-1 -
PloS One 2017Neisseria gonorrhoeae, the causative agent of gonorrhea, has a number of factors known to contribute to pathogenesis; however, a full understanding of these processes...
Neisseria gonorrhoeae, the causative agent of gonorrhea, has a number of factors known to contribute to pathogenesis; however, a full understanding of these processes and their regulation has proven to be elusive. Post-translational modifications (PTMs) of bacterial proteins are now recognized as one mechanism of protein regulation. In the present study, Western blot analyses, with an anti-acetyl-lysine antibody, indicated that a large number of gonococcal proteins are post-translationally modified. Previous work has shown that Nε-lysine acetylation can occur non-enzymatically with acetyl-phosphate (AcP) as the acetyl donor. In the current study, an acetate kinase mutant (1291ackA), which accumulates AcP, was generated in N. gonorrhoeae. Broth cultures of N. gonorrhoeae 1291wt and 1291ackA were grown, proteins extracted and digested, and peptides containing acetylated-lysines (K-acetyl) were affinity-enriched from both strains. Mass spectrometric analyses of these samples identified a total of 2686 unique acetylation sites. Label-free relative quantitation of the K-acetyl peptides derived from the ackA and wild-type (wt) strains demonstrated that 109 acetylation sites had an ackA/wt ratio>2 and p-values <0.05 in at least 2/3 of the biological replicates and were designated as "AckA-dependent". Regulated K-acetyl sites were found in ribosomal proteins, central metabolism proteins, iron acquisition and regulation proteins, pilus assembly and regulation proteins, and a two-component response regulator. Since AckA is part of a metabolic pathway, comparative growth studies of the ackA mutant and wt strains were performed. The mutant showed a growth defect under aerobic conditions, an inability to grow anaerobically, and a defect in biofilm maturation. In conclusion, the current study identified AckA-dependent acetylation sites in N. gonorrhoeae and determined that these sites are found in a diverse group of proteins. This work lays the foundation for future studies focusing on specific acetylation sites that may have relevance in gonococcal pathogenesis and metabolism.
Topics: Acetate Kinase; Acetylation; Bacterial Proteins; Gene Expression Regulation, Bacterial; Mass Spectrometry; Metabolic Networks and Pathways; Neisseria gonorrhoeae; Phosphorylation; Protein Processing, Post-Translational
PubMed: 28654654
DOI: 10.1371/journal.pone.0179621 -
Antioxidants & Redox Signaling Aug 2018Thioredoxin reductase 1 (TrxR1) is a cancer target and essential selenoprotein that defends the cell against reactive oxygen species and regulates cellular signaling and...
AIMS
Thioredoxin reductase 1 (TrxR1) is a cancer target and essential selenoprotein that defends the cell against reactive oxygen species and regulates cellular signaling and redox pathways. Previous cell-based studies correlated TrxR1 acetylation with modulated cellular reduction activity, yet the function of specific acetylation sites on TrxR1 remains unknown.
INNOVATION
We produced site-specifically acetylated TrxR1 variants that also contain selenocysteine (Sec). We demonstrated efficient high-fidelity protein synthesis with 22 different amino acids by simultaneous UAG codon reassignment to N-acetyl-lysine and UGA codon recoding to Sec.
RESULTS
We characterized TrxR1 variants acetylated at physiologically relevant sites and found that single acetylation sites increased TrxR1 activity, enhancing the apparent catalytic rate up to 2.7-fold. The activity increase in acetylated TrxR1 (acTrxR1) is reversible and is reduced following deacetylation with histone deacetylase.
CONCLUSION
Here we present a novel mechanism through which acetylation increases TrxR1 activity by destabilizing low-activity TrxR1 multimers, increasing the population of active dimeric TrxR1. Antioxid. Redox Signal. 29, 377-388.
Topics: Acetylation; Humans; Models, Molecular; Thioredoxin Reductase 1
PubMed: 29117711
DOI: 10.1089/ars.2017.7082 -
MBio Jun 2022ε-lysine acetylation is a common posttranslational modification observed in diverse species of bacteria. Aside from a few central metabolic enzymes and transcription...
ε-lysine acetylation is a common posttranslational modification observed in diverse species of bacteria. Aside from a few central metabolic enzymes and transcription factors, little is known about how this posttranslational modification regulates protein activity. In this work, we investigated how lysine acetylation affects translation in Escherichia coli. In multiple species of bacteria, ribosomal proteins are highly acetylated at conserved lysine residues, suggesting that this modification may regulate translation. In support of this hypothesis, we found that the addition of either of the acetyl donors acetyl phosphate and acetyl-coenzyme A inhibits translation but not transcription using an E. coli cell-free system. Further investigations using assays revealed that acetylation does not appear to alter the rate of translation elongation but, rather, increases the proportions of dissociated 30S and 50S ribosomes, based on polysome profiles of mutants or growth conditions known to promote lysine acetylation. Furthermore, ribosomal proteins are more acetylated in the disassociated 30S and 50S ribosomal subunits than in the fully assembled 70S complex. The effect of acetylation is also growth rate dependent, with disassociation of the subunits being most pronounced during late-exponential and early-stationary-phase growth-the same growth phase where protein acetylation is greatest. Collectively, our data demonstrate that lysine acetylation inhibits translation, most likely by interfering with subunit association. These results have also uncovered a new mechanism for coupling translation to the metabolic state of the cell. Numerous cellular processes are regulated in response to the metabolic state of the cell. One such regulatory mechanism involves lysine acetylation, a covalent modification involving the transfer of an acetyl group from central metabolite acetyl-coenzyme A or acetyl phosphate to a lysine residue in a protein. This posttranslational modification is known to regulate some central metabolic enzymes and transcription factors in bacteria, though a comprehensive understanding of its effect on cellular physiology is still lacking. In the present study, lysine acetylation was also found to inhibit translation in Escherichia coli by impeding ribosome association, most likely by disrupting salt bridges along the binding interface of the 30S and 50S ribosomal subunits. These results further our understanding of lysine acetylation by uncovering protein synthesis as a new target of regulation and aid in the design of bacteria for biotechnology applications where the growth conditions are known to promote lysine acetylation.
Topics: Acetyl Coenzyme A; Acetylation; Escherichia coli; Lysine; Protein Processing, Post-Translational; Ribosomal Proteins; Transcription Factors
PubMed: 35604121
DOI: 10.1128/mbio.01224-22 -
The Plant Journal : For Cell and... May 2023Polyamines such as spermidine and spermine are essential regulators of cell growth, differentiation, maintenance of ion balance and abiotic stress tolerance. Their...
Polyamines such as spermidine and spermine are essential regulators of cell growth, differentiation, maintenance of ion balance and abiotic stress tolerance. Their levels are controlled by the spermidine/spermine N -acetyltransferase (SSAT) via acetylation to promote either their degradation or export outside the cell as shown in mammals. Plant genomes contain at least one gene coding for SSAT (also named NATA for N-AcetylTransferase Activity). Combining kinetics, HPLC-MS and crystallography, we show that three plant SSATs, one from the lower plant moss Physcomitrium patens and two from the higher plant Zea mays, acetylate various aliphatic polyamines and two amino acids lysine (Lys) and ornithine (Orn). Thus, plant SSATs exhibit a broad substrate specificity, unlike more specific human SSATs (hSSATs) as hSSAT1 targets polyamines, whereas hSSAT2 acetylates Lys and thiaLys. The crystal structures of two PpSSAT ternary complexes, one with Lys and CoA, the other with acetyl-CoA and polyethylene glycol (mimicking spermine), reveal a different binding mode for polyamine versus amino acid substrates accompanied by structural rearrangements of both the coenzyme and the enzyme. Two arginine residues, unique among plant SSATs, hold the carboxyl group of amino acid substrates. The most abundant acetylated compound accumulated in moss was N -acetyl-Lys, whereas N -acetyl-Orn, known to be toxic for aphids, was found in maize. Both plant species contain very low levels of acetylated polyamines. The present study provides a detailed biochemical and structural basis of plant SSAT enzymes that can acetylate a wide range of substrates and likely play various roles in planta.
Topics: Animals; Humans; Polyamines; Spermidine; Spermine; Zea mays; Lysine; Ornithine; Acetylation; Acetyltransferases; Catalysis; Mammals
PubMed: 36786691
DOI: 10.1111/tpj.16148 -
Cell Death and Differentiation Apr 2006Gradual disclosure of the molecular basis of selective neuronal apoptosis during neurodegenerative diseases reveals active participation of acetylating and deacetylating... (Review)
Review
Gradual disclosure of the molecular basis of selective neuronal apoptosis during neurodegenerative diseases reveals active participation of acetylating and deacetylating agents during the process. Several studies have now successfully manipulated neuronal vulnerability by influencing the dose and enzymatic activity of histone acetyltransferases (HATs) and histone deacetylases (HDACs), enzymes regulating acetylation homeostasis within the nucleus, thus focusing on the importance of balanced acetylation status in neuronal vitality. It is now increasingly becoming clear that acetylation balance is greatly impaired during neurodegenerative conditions. Herein, we attempt to illuminate molecular means by which such impairment is manifested and how the compromised acetylation homeostasis is intimately coupled to neurodegeneration. Finally, we discuss the therapeutic potential of reinstating the HAT-HDAC balance to ameliorate neurodegenerative diseases.
Topics: Acetylation; Animals; Apoptosis; Butyrates; Enzyme Inhibitors; Histone Acetyltransferases; Histone Deacetylase Inhibitors; Histone Deacetylases; Homeostasis; Humans; Neurodegenerative Diseases; Neurons; Signal Transduction
PubMed: 16167067
DOI: 10.1038/sj.cdd.4401769 -
Molecular & Cellular Proteomics : MCP Jul 2002The acetylation isoforms of histone H4 from butyrate-treated HeLa cells were separated by C(4) reverse-phase high pressure liquid chromatography and by polyacrylamide...
The acetylation isoforms of histone H4 from butyrate-treated HeLa cells were separated by C(4) reverse-phase high pressure liquid chromatography and by polyacrylamide gel electrophoresis. Histone H4 bands were excised and digested in-gel with the endoprotease trypsin. Matrix-assisted laser desorption ionization time-of-flight mass spectrometry was used to characterize the level of acetylation, and nanoelectrospray tandem mass spectrometric analysis of the acetylated peptides was used to determine the exact sites of acetylation. Although there are 15 acetylation sites possible, only four acetylated peptide sequences were actually observed. The tetra-acetylated form is modified at lysines 5, 8, 12, and 16, the tri-acetylated form is modified at lysines 8, 12, and 16, and the di-acetylated form is modified at lysines 12 and 16. The only significant amount of the mono-acetylated form was found at position 16. These results are consistent with the hypothesis of a "zip" model whereby acetylation of histone H4 proceeds in the direction of from Lys-16 to Lys-5, and deacetylation proceeds in the reverse direction. Histone acetylation and deacetylation are coordinated processes leading to a non-random distribution of isoforms. Our results also revealed that lysine 20 is di-methylated in all modified isoforms, as well as the non-acetylated isoform of H4.
Topics: Acetylation; HeLa Cells; Histones; Humans; Models, Biological; Peptide Fragments; Spectrometry, Mass, Electrospray Ionization; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
PubMed: 12239278
DOI: 10.1074/mcp.m200031-mcp200 -
The Biochemical Journal Oct 2018Deciphering the histone code has illustrated that acetylation or methylation on the same residue can have analogous or opposing roles. However, little is known about the...
Deciphering the histone code has illustrated that acetylation or methylation on the same residue can have analogous or opposing roles. However, little is known about the interplay between these post-translational modifications (PTMs) on the same nonhistone residues. We have recently discovered that N-terminal acetyltransferases (NATs) and N-terminal methyltransferases (NRMTs) can have overlapping substrates and identified myosin regulatory light chain 9 (MYL9) as the first confirmed protein to occur in either α-amino-methylated (Nα-methyl) or α-amino-acetylated (Nα-acetyl) states Here we aim to determine if these PTMs function similarly or create different MYL9 proteoforms with distinct roles. We use enzymatic assays to directly verify MYL9 is a substrate of both NRMT1 and NatA and generate mutants of MYL9 that are exclusive for Nα-acetylation or Nα-methylation. We then employ eukaryotic cell models to probe the regulatory functions of these Nα-PTMs on MYL9. Our results show that, contrary to prevailing dogma, neither of these modifications regulate the stability of MYL9. Rather, exclusive Nα-acetylation promotes cytoplasmic roles of MYL9, while exclusive Nα-methylation promotes the nuclear role of MYL9 as a transcription factor. The increased cytoplasmic activity of Nα-acetylated MYL9 corresponds with increased phosphorylation at serine 19, a key MYL9 activating PTM. Increased nuclear activity of Nα-methylated MYL9 corresponds with increased DNA binding. Nα-methylation also results in a decrease of interactions between the N-terminus of MYL9 and a host of cytoskeletal proteins. These results confirm that Nα-acetylation and Nα-methylation differentially affect MYL9 function by creating distinct proteoforms with different internal PTM patterns and binding properties.
Topics: Acetylation; Animals; Cell Movement; HCT116 Cells; HEK293 Cells; Humans; Methylation; Mice; Myosin Light Chains; NIH 3T3 Cells
PubMed: 30242065
DOI: 10.1042/BCJ20180638 -
Scientific Reports Mar 2017ATP synthases (ATPases) are enzymes that produce ATP and control the pH in the cell or cellular compartments. While highly conserved over different species, ATPases are...
ATP synthases (ATPases) are enzymes that produce ATP and control the pH in the cell or cellular compartments. While highly conserved over different species, ATPases are structurally well-characterised but the existence and functional significance of many post-translational modifications (PTMs) is not well understood. We combined a range of mass spectrometric techniques to unravel the location and extent of PTMs in the chloroplast ATP synthase (cATPase) purified from spinach leaves. We identified multiple phosphorylation and acetylation sites and found that both modifications stabilise binding of ε and δ subunits. Comparing cross-linking of naturally modified cATPase with the in vitro deacetylated enzyme revealed a major conformational change in the ε subunit in accord with extended and folded forms of the subunit. Locating modified residues within the catalytic head we found that phosphorylated and acetylated residues are primarily on α/β and β/α interfaces respectively. By aligning along different interfaces the higher abundance acetylated residues are proximal to the regulatory sites while the lower abundance phosphorylation sites are more densely populated at the catalytic sites. We propose that modifications in the catalytic head, together with the conformational change in subunit ε, work in synergy to fine-tune the enzyme during adverse conditions.
Topics: Acetylation; Chloroplast Proteins; Enzyme Stability; Phosphorylation; Proton-Translocating ATPases
PubMed: 28276484
DOI: 10.1038/srep44068