scholarly journals Differential expression of 5-enol-pyruvyl-shikimate-3-phosphate synthase isoforms in elicitor-treated, cultured maize cells

2002 ◽  
Vol 29 (12) ◽  
pp. 1483 ◽  
Author(s):  
Giuseppe Forlani
Keyword(s):  
Phenylalanine Ammonia Lyase ◽  
Zea Mays L ◽  
Cultured Cells ◽  
Amino Acid Content ◽  
Shikimate Pathway ◽  
Activity Levels ◽  
Sharp Rise ◽  
Enzyme Form ◽  
Genes Encoding ◽  
Phosphate Synthase

The expression of two 5-enol-pyruvyl-shikimate-3-phosphate synthase (EC 2.5.1.19) isoforms was investigated in Zea mays L. suspension-cultured cells following exposure to a fungal elicitor. Activity levels of isozyme II specifically increased soon after treatment, in strict connection with induction of phenylalanine ammonia-lyase (PAL) and attainment of a new free-phenylalanine homeostasis at a higher concentration. However, a few days later, activity of the other enzyme form was also significantly enhanced, concomitant with a sharp rise in overall amino acid content, a further increase in PAL level and a resumption of cell lysis. Besides strengthening the hypothesis that an entire set of genes encoding for shikimate pathway enzymes (whose expression is specifically involved in plant dynamic defence) may exist, a general change in the levels of several amino acids seems to point towards a reprogramming of their metabolism in elicited cells.

Differential expression of tomato (Lycopersicon esculentum L.) genes encoding shikimate pathway isoenzymes. I. 3-Deoxy-D-arabino-heptulosonate 7-phosphate synthase

1993 ◽  
Vol 23 (4) ◽  
pp. 697-706 ◽  
Author(s):  
J�rn G�rlach ◽  
Andreas Beck ◽  
John M. Henstrand ◽  
Avtar K. Handa ◽  
Klaus M. Herrmann ◽  
...  
Keyword(s):  
Lycopersicon Esculentum ◽  
Differential Expression ◽  
Shikimate Pathway ◽  
Genes Encoding ◽  
Phosphate Synthase

scholarly journals β-Galactosidases from a Sequence-Based Metagenome: Cloning, Expression, Purification and Characterization

2020 ◽  
Vol 9 (1) ◽  
pp. 55
Author(s):  
María Florencia Eberhardt ◽  
José Matías Irazoqui ◽  
Ariel Fernando Amadio
Keyword(s):  
Bacterial Species ◽  
Value Added ◽  
Raw Material ◽  
Activity Levels ◽  
Treatment Technology ◽  
Catalytic Domains ◽  
Stabilization Ponds ◽  
Cazy Database ◽  
Genes Encoding ◽  
Value Added Products

Stabilization ponds are a common treatment technology for wastewater generated by dairy industries. Large proportions of cheese whey are thrown into these ponds, creating an environmental problem because of the large volume produced and the high biological and chemical oxygen demands. Due to its composition, mainly lactose and proteins, it can be considered as a raw material for value-added products, through physicochemical or enzymatic treatments. β-Galactosidases (EC 3.2.1.23) are lactose modifying enzymes that can transform lactose in free monomers, glucose and galactose, or galactooligosacharides. Here, the identification of novel genes encoding β-galactosidases, identified via whole-genome shotgun sequencing of the metagenome of dairy industries stabilization ponds is reported. The genes were selected based on the conservation of catalytic domains, comparing against the CAZy database, and focusing on families with β-galactosidases activity (GH1, GH2 and GH42). A total of 394 candidate genes were found, all belonging to bacterial species. From these candidates, 12 were selected to be cloned and expressed. A total of six enzymes were expressed, and five cleaved efficiently ortho-nitrophenyl-β-galactoside and lactose. The activity levels of one of these novel β-galactosidase was higher than other enzymes reported from functional metagenomics screening and higher than the only enzyme reported from sequence-based metagenomics. A group of novel mesophilic β-galactosidases from diary stabilization ponds’ metagenomes was successfully identified, cloned and expressed. These novel enzymes provide alternatives for the production of value-added products from dairy industries’ by-products.

scholarly journals Structure-guided selection of puromycin N-acetyltransferase mutants with enhanced selection stringency for deriving mammalian cell lines expressing recombinant proteins

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Alessandro T. Caputo ◽  
Oliver M. Eder ◽  
Hana Bereznakova ◽  
Heleen Pothuis ◽  
Albert Ardevol ◽  
...  
Keyword(s):  
Cell Lines ◽  
Mammalian Cell ◽  
Recombinant Proteins ◽  
Cultured Cells ◽  
Hek293 Cells ◽  
Reaction Products ◽  
Enzyme Form ◽  
X Ray Crystallography ◽  
Mammalian Cell Lines ◽  
Apparent Loss

AbstractPuromycin and the Streptomyces alboniger-derived puromycin N-acetyltransferase (PAC) enzyme form a commonly used system for selecting stably transfected cultured cells. The crystal structure of PAC has been solved using X-ray crystallography, revealing it to be a member of the GCN5-related N-acetyltransferase (GNAT) family of acetyltransferases. Based on structures in complex with acetyl-CoA or the reaction products CoA and acetylated puromycin, four classes of mutations in and around the catalytic site were designed and tested for activity. Single-residue mutations were identified that displayed a range of enzymatic activities, from complete ablation to enhanced activity relative to wild-type (WT) PAC. Cell pools of stably transfected HEK293 cells derived using two PAC mutants with attenuated activity, Y30F and A142D, were found to secrete up to three-fold higher levels of a soluble, recombinant target protein than corresponding pools derived with the WT enzyme. A third mutant, Y171F, appeared to stabilise the intracellular turnover of PAC, resulting in an apparent loss of selection stringency. Our results indicate that the structure-guided manipulation of PAC function can be utilised to enhance selection stringency for the derivation of mammalian cell lines secreting elevated levels of recombinant proteins.

scholarly journals Transcription Profiles Analysis of Genes Encoding 1-Deoxy-D-xylulose 5-Phosphate Synthase and 2C-Methyl-D-erythritol 4-Phosphate Synthase in Plaunotol Biosynthesis from Croton stellatopilosus

2008 ◽  
Vol 31 (5) ◽  
pp. 852-856 ◽  
Author(s):  
Juraithip Wungsintaweekul ◽  
Tanawan Sirisuntipong ◽  
Damrong Kongduang ◽  
Thanaisawan Losuphanporn ◽  
Anan Ounaroon ◽  
...  
Keyword(s):  
Transcription Profiles ◽  
Genes Encoding ◽  
Phosphate Synthase

scholarly journals Identification of Structural Variants in Two Novel Genomes of Maize Inbred Lines Possibly Related to Glyphosate Tolerance

Plants ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 523
Author(s):  
Medhat Mahmoud ◽  
Joanna Gracz-Bernaciak ◽  
Marek Żywicki ◽  
Wojciech Karłowski ◽  
Tomasz Twardowski ◽  
...  
Keyword(s):  
Gene Expression ◽  
Single Molecule ◽  
Zea Mays L ◽  
Shikimate Pathway ◽  
High Impact ◽  
Maize Genome ◽  
Structural Variants ◽  
Shikimate Dehydrogenase ◽  
Epsps Gene ◽  
Sequencing Technologies

To study genetic variations between genomes of plants that are naturally tolerant and sensitive to glyphosate, we used two Zea mays L. lines traditionally bred in Poland. To overcome the complexity of the maize genome, two sequencing technologies were employed: Illumina and Single Molecule Real-Time (SMRT) PacBio. Eleven thousand structural variants, 4 million SNPs and approximately 800 thousand indels differentiating the two genomes were identified. Detailed analyses allowed to identify 20 variations within the EPSPS gene, but all of them were predicted to have moderate or unknown effects on gene expression. Other genes of the shikimate pathway encoding bifunctional 3-dehydroquinate dehydratase/shikimate dehydrogenase and chorismate synthase were altered by variants predicted to have a high impact on gene expression. Additionally, high-impact variants located within the genes involved in the active transport of glyphosate through the cell membrane encoding phosphate transporters as well as multidrug and toxic compound extrusion have been identified.

The nuclear receptor for bile acids, FXR, transactivates human organic solute transporter-α and -β genes

2006 ◽  
Vol 290 (3) ◽  
pp. G476-G485 ◽  
Author(s):  
Jean-François Landrier ◽  
Jyrki J. Eloranta ◽  
Stephan R. Vavricka ◽  
Gerd A. Kullak-Ublick
Keyword(s):  
Bile Acid ◽  
Bile Acids ◽  
Cultured Cells ◽  
Farnesoid X Receptor ◽  
Mrna Levels ◽  
Small Interfering Rnas ◽  
Organic Solute Transporter ◽  
Genes Encoding ◽  
Organic Solute ◽  
Solute Transporter

Bile acids are synthesized from cholesterol in the liver and are excreted into bile via the hepatocyte canalicular bile salt export pump. After their passage into the intestine, bile acids are reabsorbed in the ileum by sodium-dependent uptake across the apical membrane of enterocytes. At the basolateral domain of ileal enterocytes, bile acids are extruded into portal blood by the heterodimeric organic solute transporter OSTα/OSTβ. Although the transport function of OSTα/OSTβ has been characterized, little is known about the regulation of its expression. We show here that human OSTα/OSTβ expression is induced by bile acids through ligand-dependent transactivation of both OST genes by the nuclear bile acid receptor/farnesoid X receptor (FXR). FXR agonists induced endogenous mRNA levels of OSTα and OSTβ in cultured cells, an effect that was not discernible upon inhibition of FXR expression by small interfering RNAs. Furthermore, OST mRNAs were induced in human ileal biopsies exposed to the bile acid chenodeoxycholic acid. Reporter constructs containing OSTα or OSTβ promoters were transactivated by FXR in the presence of its ligand. Two functional FXR binding motifs were identified in the OSTα gene and one in the OSTβ gene. Targeted mutation of these elements led to reduced inducibility of both OST promoters by FXR. In conclusion, the genes encoding the human OSTα/OSTβ complex are induced by bile acids and FXR. By coordinated control of OSTα/OSTβ expression, bile acids may adjust the rate of their own efflux from enterocytes in response to changes in intracellular bile acid levels.

scholarly journals Multiple Rare Risk Coding Variants in Postsynaptic Density-Related Genes Associated With Schizophrenia Susceptibility

2020 ◽  
Vol 11 ◽  
Author(s):  
Tsung-Ming Hu ◽  
Ying-Chieh Wang ◽  
Chia-Liang Wu ◽  
Shih-Hsin Hsu ◽  
Hsin-Yao Tsai ◽  
...  
Keyword(s):  
Protein Function ◽  
Cultured Cells ◽  
Postsynaptic Density ◽  
Psychiatric History ◽  
Missense Mutations ◽  
Genes Encoding ◽  
Schizophrenic Patients ◽  
Synaptic Receptors ◽  
Related Proteins ◽  
Coding Variants

ObjectiveSchizophrenia is a chronic debilitating neurobiological disorder of aberrant synaptic connectivity and synaptogenesis. Postsynaptic density (PSD)–related proteins in N-methyl-D-aspartate receptor–postsynaptic signaling complexes are crucial to regulating the synaptic transmission and functions of various synaptic receptors. This study examined the role of PSD-related genes in susceptibility to schizophrenia.MethodsWe resequenced 18 genes encoding the disks large-associated protein (DLGAP), HOMER, neuroligin (NLGN), neurexin, and SH3 and multiple ankyrin repeat domains (SHANK) protein families in 98 schizophrenic patients with family psychiatric history using semiconductor sequencing. We analyzed the protein function of the identified rare schizophrenia-associated mutants via immunoblotting and immunocytochemistry.ResultsWe identified 50 missense heterozygous mutations in 98 schizophrenic patients with family psychiatric history, and in silico analysis revealed some as damaging or pathological to the protein function. Ten missense mutations were absent from the dbSNP database, the gnomAD (non-neuro) dataset, and 1,517 healthy controls from Taiwan BioBank. Immunoblotting revealed eight missense mutants with altered protein expressions in cultured cells compared with the wild type.ConclusionOur findings suggest that PSD-related genes, especially the NLGN, SHANK, and DLGAP families, harbor rare functional mutations that might alter protein expression in some patients with schizophrenia, supporting contributing rare coding variants into the genetic architecture of schizophrenia.

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