scholarly journals Human HemK2/KMT9/N6AMT1 is an active protein methyltransferase, but does not act on DNA in vitro, in the presence of Trm112

2019 ◽  
Vol 5 (1) ◽  
Author(s):  
Clayton B. Woodcock ◽  
Dan Yu ◽  
Xing Zhang ◽  
Xiaodong Cheng
Keyword(s):  
Active Protein ◽  
Protein Methyltransferase

Half-Life of Platelet Factor 4 (PF-4) in Plasma and Platelets from Macaca Mulatta

1977 ◽  
Vol 37 (01) ◽  
pp. 073-080 ◽  
Author(s):  
Knut Gjesdal ◽  
Duncan S. Pepper
Keyword(s):  
Macaca Mulatta ◽  
Half Life ◽  
Human Platelet ◽  
Gel Filtration ◽  
Platelet Factor 4 ◽  
Active Protein ◽  
Platelet Factor ◽  
Membrane Bound

SummaryHuman platelet factor 4 (PF-4) showed a reaction of complete identity with PF-4 from Macaca mulatta when tested against rabbit anti-human-PF-4. Such immunoglobulin was used for quantitative precipitation of in vivo labelled PF-4 in monkey serum. The results suggest that the active protein had an intra-platelet half-life of about 21 hours. In vitro 125I-labelled human PF-4 was injected intravenously into two monkeys and isolated by immuno-precipita-tion from platelet-poor plasma and from platelets disrupted after gel-filtration. Plasma PF-4 was found to have a half-life of 7 to 11 hours. Some of the labelled PF-4 was associated with platelets and this fraction had a rapid initial disappearance rate and a subsequent half-life close to that of plasma PF-4. The results are compatible with the hypothesis that granular PF-4 belongs to a separate compartment, whereas membrane-bound PF-4 and plasma PF-4 may interchange.

scholarly journals SET7/9-dependent methylation of ARTD1 at K508 stimulates poly-ADP-ribose formation after oxidative stress

Open Biology ◽  
2013 ◽  
Vol 3 (10) ◽  
pp. 120173 ◽  
Author(s):  
Ingrid Kassner ◽  
Anneli Andersson ◽  
Monika Fey ◽  
Martin Tomas ◽  
Elisa Ferrando-May ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Protein Interactions ◽  
Protein Function ◽  
Dependent Manner ◽  
Protein Protein Interactions ◽  
Post Translational Modification ◽  
Target Proteins ◽  
Protein Methyltransferase

ADP-ribosyltransferase diphtheria toxin-like 1 (ARTD1, formerly PARP1) is localized in the nucleus, where it ADP-ribosylates specific target proteins. The post-translational modification (PTM) with a single ADP-ribose unit or with polymeric ADP-ribose (PAR) chains regulates protein function as well as protein–protein interactions and is implicated in many biological processes and diseases. SET7/9 (Setd7, KMT7) is a protein methyltransferase that catalyses lysine monomethylation of histones, but also methylates many non-histone target proteins such as p53 or DNMT1. Here, we identify ARTD1 as a new SET7/9 target protein that is methylated at K508 in vitro and in vivo . ARTD1 auto-modification inhibits its methylation by SET7/9, while auto-poly-ADP-ribosylation is not impaired by prior methylation of ARTD1. Moreover, ARTD1 methylation by SET7/9 enhances the synthesis of PAR upon oxidative stress in vivo . Furthermore, laser irradiation-induced PAR formation and ARTD1 recruitment to sites of DNA damage in a SET7/9-dependent manner. Together, these results reveal a novel mechanism for the regulation of cellular ARTD1 activity by SET7/9 to assure efficient PAR formation upon cellular stress.

scholarly journals Characterization of prenylated protein methyltransferase in Leishmania

1999 ◽  
Vol 342 (3) ◽  
pp. 513-518 ◽  
Author(s):  
Marie-Pierre HASNE ◽  
Françoise LAWRENCE
Keyword(s):  
Protein Kinase C ◽  
Leishmania Donovani ◽  
Post Translational Modification ◽  
Methyltransferase Activity ◽  
Kinase C ◽  
Protein Methyltransferase ◽  
Molecular Masses ◽  
Endogenous Substrates

Prenylated protein methyltransferase, an enzyme involved in the post-translational modification of many signalling proteins, has been characterized in a parasitic flagellated protozoan, Leishmania donovani. The activity of this enzyme was monitored by the methylation of an artificial substrate, an S-prenylated cysteine analogue, with S-adenosyl-L-[methyl-3H]methionine as methyl donor. More than 85% of the methyltransferase activity was associated with membranes. The enzyme methylates N-acetyl-S-trans,trans-farnesyl-L-cysteine and N-acetyl-S-all-trans-geranylgeranyl-L-cysteine, but N-acetyl-S-trans,trans-geranyl-L-cysteine only very weakly. In contrast with the enzyme from mammals, the leishmanial enzyme had a greater affinity for the farnesylated substrate than for the geranylgeranylated one. Activity in vitro was not modulated by cAMP, protein kinase C activator or guanosine 5′-[γ-thio]triphosphate. An analysis of the endogenous substrates showed that the carboxymethylated proteins were also isoprenylated. The main carboxymethylated proteins have molecular masses of 95, 68, 55, 46, 34-23, 18 and less than 14 kDa. Treatment of cells with N-acetyl-S-trans,trans-farnesyl-L-cysteine decreased the carboxymethylation level, whereas treatment with guanosine 5′-[γ-thio]triphosphate increased the carboxymethylation of various proteins, particularly those of molecular masses 30-20 kDa.

scholarly journals SUPPRESSION OF DELAYED HYPERSENSITIVITY IN VITRO BY INHIBITION OF PROTEIN SYNTHESIS

1965 ◽  
Vol 122 (6) ◽  
pp. 1125-1134 ◽  
Author(s):  
John R. David
Keyword(s):  
Protein Synthesis ◽  
Culture Medium ◽  
Specific Antigen ◽  
Delayed Hypersensitivity ◽  
Cell Protein ◽  
Sensitive Cell ◽  
Active Protein ◽  
Inhibit Protein Synthesis ◽  
Inhibition Of Protein Synthesis

Peritoneal cells from guinea pigs exhibiting delayed hypersensitivity are inhibited from migrating in vitro by specific antigen. This inhibition is prevented by the addition of puromycin to the culture medium. The amount of puromycin necessary to prevent the inhibition by antigen also suppressed the incorporation of C14-leucine into peritoneal cell protein. Additional evidence that the action of puromycin is due to its inhibition of protein synthesis has been obtained with analogues of puromycin; those that inhibit protein synthesis also prevent the action of antigen on the cells, while those analogues that do not inhibit protein synthesis have no effect. Actinomycin also prevents the inhibition of sensitive cells by antigen while chloramphenicol has no effect. The data indicate that the inhibition of sensitive cell migration by antigen requires active protein synthesis. The possible mechanisms by which inhibition of protein synthesis may influence the in vitro reactions of delayed hypersensitivity are discussed.

scholarly journals 1,N6-α-hydroxypropanoadenine, the acrolein adduct to adenine, is a substrate for AlkB dioxygenase

2017 ◽  
Vol 474 (11) ◽  
pp. 1837-1852 ◽  
Author(s):  
Małgorzata Dylewska ◽  
Jarosław T. Kuśmierek ◽  
Tomasz Pilżys ◽  
Jarosław Poznański ◽  
Agnieszka M. Maciejewska
Keyword(s):  
Substrate Binding ◽  
Protonated Form ◽  
Environmental Pollutant ◽  
Hydroxyl Group ◽  
Active Protein ◽  
Biochemical Activity ◽  
Differential Scanning Fluorimetry ◽  
Reaction Cycles

1,N6-α-hydroxypropanoadenine (HPA) is an exocyclic DNA adduct of acrolein – an environmental pollutant and endocellular oxidative stress product. Escherichia coli AlkB dioxygenase belongs to the superfamily of α-ketoglutarate (αKG)- and iron-dependent dioxygenases which remove alkyl lesions from bases via an oxidative mechanism, thereby restoring native DNA structure. Here, we provide in vivo and in vitro evidence that HPA is mutagenic and is effectively repaired by AlkB dioxygenase. HPA generated in plasmid DNA caused A → C and A → T transversions and, less frequently, A → G transitions. The lesion was efficiently repaired by purified AlkB protein; the optimal pH, Fe(II), and αKG concentrations for this reaction were determined. In vitro kinetic data show that the protonated form of HPA is preferentially repaired by AlkB, albeit the reaction is stereoselective. Moreover, the number of reaction cycles carried out by an AlkB molecule remains limited. Molecular modeling of the T(HPA)T/AlkB complex demonstrated that the R stereoisomer in the equatorial conformation of the HPA hydroxyl group is strongly preferred, while the S stereoisomer seems to be susceptible to AlkB-directed oxidative hydroxylation only when HPA adopts the syn conformation around the glycosidic bond. In addition to the biochemical activity assays, substrate binding to the protein was monitored by differential scanning fluorimetry allowing identification of the active protein form, with cofactor and cosubstrate bound, and monitoring of substrate binding. In contrast FTO, a human AlkB homolog, failed to bind an ssDNA trimer carrying HPA.

Changes in Ribosome Organization and Messenger RNA Abundance in Ripening Tomato Fruits

1984 ◽  
Vol 11 (3) ◽  
pp. 225 ◽  
Author(s):  
J Spiers ◽  
CJ Brady ◽  
D Grierson ◽  
E Lee
Keyword(s):  
Protein Synthesis ◽  
Ribosomal Rna ◽  
Messenger Rna ◽  
In Vitro Translation ◽  
Translation System ◽  
Active Protein ◽  
Tomato Fruits ◽  
Rna And Protein Synthesis ◽  
Early Ripening

The involvement of RNA and protein synthesis in fruit ripening was investigated. Mature-green tomato fruits were found to contain about 30% of their ribosomal RNA in polyribosomes. At the 'breaker' or early ripening stage, about 50% of rRNA was in polyribosomes and this distribution of rRNA was maintained until fruits were fully ripe. The continued presence of polyribosomes is consistent with active protein synthesis persisting through and beyond the climacteric period when the wall-hydrolysing enzyme polygalacturonase accumulates and fruits soften, synthesize lycopene and undergo other ripening related changes. Poly(A)-containing RNA purified from polyribosomes extracted from individual fruits was used to prime the synthesis of [35S]methionine-labelled polypeptides by a wheat germ in vitro translation system. The pattern of polypeptides synthesized in response to RNA from mature-green fruits differed from that given by RNA from ripening fruits. The majority of changes were found to occur within approximately 48 h of the increase of ethylene synthesis and were apparent in all fruits with any pink or red colour. Similar results were obtained by translating total cellular RNA and total polyribosomal RNA indicating that the major RNA species shown in this study to change in abundance during ripening are polyadenylated, and hence most probably cytoplasmic, and do not accumulate as 'stored messages' outside of the polyribosomes. The differences between green and ripening fruits in polyribosome profiles were demonstrated in two cultivars of tomato. Differences in mRNA populations between green and ripe fruits were found in three cultivars.

ON THE MECHANISM OF HORMONE ACTION. VII.

1968 ◽  
Vol 57 (1) ◽  
pp. 33-44 ◽  
Author(s):  
N. Lang ◽  
P. Herrlich ◽  
C. E. Sekeris
Keyword(s):  
Protein Synthesis ◽  
Enzyme Activity ◽  
Messenger Rna ◽  
De Novo ◽  
Hormone Action ◽  
Transaminase Activity ◽  
Active Protein ◽  
Tyrosine Transaminase ◽  
Mechanism Of Hormone Action

ABSTRACT A messenger RNA fraction has been isolated from the liver of cortisol treated rats which when added to an in vitro protein synthesizing sytem gives rise to tyrosine-α-ketoglutarate transaminase activity. Development of enzyme activity is dependent on active protein synthesis and is inhibited by puromycin and erythromycin. It is concluded that the messenger RNA fraction contains the information for the formation of tyrosine transaminase and that the enzyme is formed in vitro by de novo synthesis.

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