Search results for "Cofactor"
showing 10 items of 74 documents
Purification and characterization of geranyl diphosphate synthase from Vitis vinifera L. cv Muscat de Frontignant cell cultures
1993
A geranyl diphosphate synthase (EC 2.5.1.1), which catalyzes the formation of geranyl diphosphate from dimethylallyl diphosphate and isopentenyl diphosphate, was isolated from Vitis vinifera L. cv Muscat de Frontignan cell cultures. Purification of the enzyme was achieved successively by ammonium sulfate precipitation and chromatography on DEAE-Sephacel, hydroxylapatite, Mono Q, Phenyl Superose, Superose 12, and preparative nondenaturing polyacrylamide gels. The enzyme formed only geranyl diphosphate as a product. In all cases, neither neryl diphosphate, the cis isomer, nor farnesyl diphosphate was detected. The enzyme showed a native molecular mass of 68 [plus or minus] 5 kD as determined …
Techniques to Analyze sRNA Protein Cofactor Self-Assembly In Vitro
2018
Post-transcriptional control of gene expression by small regulatory noncoding RNA (sRNA) needs protein accomplices to occur. Past research mainly focused on the RNA chaperone Hfq as cofactor. Nevertheless, recent studies indicated that other proteins might be involved in sRNA-based regulations. As some of these proteins have been shown to self-assemble, we describe in this chapter protocols to analyze the nano-assemblies formed. Precisely, we focus our analysis on Escherichia coli Hfq as a model, but the protocols presented here can be applied to analyze any polymer of proteins. This chapter thus provides a guideline to develop commonly used approaches to detect prokaryotic protein self-ass…
The Expression of NOX From Synthetic Promoters Reveals an Important Role of the Redox Status in Regulating Secondary Metabolism of
2020
Redox cofactors play a pivotal role in primary cellular metabolism, whereas the clear link between redox status and secondary metabolism is still vague. In this study we investigated effects of redox perturbation on the production of erythromycin in Saccharopolyspora erythraea by expressing the water-forming NADH oxidase (NOX) from Streptococcus pneumonia at different levels with synthetic promoters. The expression of NOX reduced the intracellular [NADH]/[NAD+] ratio significantly in S. erythraea which resulted in an increased production of erythromycin by 19∼29% and this increment rose to 60% as more oxygen was supplied. In contrast, the lower redox ratio resulted in a decreased production…
Depleted uranium induces human carcinogenesis involving the immune and chaperoning systems: Realities and working hypotheses
2019
Abstract Cancer is caused by a combination of factors, genetic, epigenetics and environmental. Among the latter, environmental pollutants absorbed by contact, inhalation, or ingestion are major proven or suspected culprits. Depleted uranium (DU) is one of them directly pertinent to the military and civilians working in militarized areas. It is considered a weak carcinogen but its implication in cancer development in exposed individuals is supported by various data. Since not all subjects exposed to DU develop cancer, it is likely that DU-dependent carcinogenesis requires cofactors, such as genetic predisposition and deficiencies of the chaperoning and immune systems. It is of the essence to…
Molecular strategies to increase yeast iron accumulation and resistance.
2018
All eukaryotic organisms rely on iron as an essential micronutrient for life because it participates as a redox-active cofactor in multiple biological processes. However, excess iron can generate reactive oxygen species that damage cellular macromolecules. The low solubility of ferric iron at physiological conditions increases the prevalence of iron deficiency anemia. A common strategy to treat iron deficiency consists of dietary iron supplementation. The baker’s yeast Saccharomyces cerevisiae is used as a model eukaryotic organism, but also as a feed supplement. In response to iron deficiency, the yeast Aft1 transcription factor activates cellular iron acquisition. However, when constituti…
Structural Analysis of Phosphoserine Aminotransferase (Isoform 1) From Arabidopsis thaliana– the Enzyme Involved in the Phosphorylated Pathway of Ser…
2018
Phosphoserine aminotransferase (PSAT) is a pyridoxal 5'-phosphate (PLP)-dependent enzyme that catalyzes the conversion of 3-phosphohydroxypyruvate (3-PHP) to 3-phosphoserine (PSer) in an L-glutamate (Glu)-linked reversible transamination reaction. This process proceeds through a bimolecular ping-pong mechanism and in plants takes place in plastids. It is a part of the phosphorylated pathway of serine biosynthesis, one of three routes recognized in plant organisms that yield serine. In this three-step biotransformation, 3-phosphoglycerate (3-PGA) delivered from plastidial glycolysis and Calvin cycle is oxidized by 3-PGA dehydrogenase. Then, 3-PHP is subjected to transamination with Glu to yi…
Characterization of sulfhydryl oxidase from Aspergillus tubingensis
2017
Background Despite of the presence of sulfhydryl oxidases (SOXs) in the secretomes of industrially relevant organisms and their many potential applications, only few of these enzymes have been biochemically characterized. In addition, basic functions of most of the SOX enzymes reported so far are not fully understood. In particular, the physiological role of secreted fungal SOXs is unclear. Results The recently identified SOX from Aspergillus tubingensis (AtSOX) was produced, purified and characterized in the present work. AtSOX had a pH optimum of 6.5, and showed a good pH stability retaining more than 80% of the initial activity in a pH range 4-8.5 within 20 h. More than 70% of the initia…
Identification and structural characterization of LytU, a unique peptidoglycan endopeptidase from the lysostaphin family
2017
AbstractWe introduce LytU, a short member of the lysostaphin family of zinc-dependent pentaglycine endopeptidases. It is a potential antimicrobial agent for S. aureus infections and its gene transcription is highly upregulated upon antibiotic treatments along with other genes involved in cell wall synthesis. We found this enzyme to be responsible for the opening of the cell wall peptidoglycan layer during cell divisions in S. aureus. LytU is anchored in the plasma membrane with the active part residing in the periplasmic space. It has a unique Ile/Lys insertion at position 151 that resides in the catalytic site-neighbouring loop and is vital for the enzymatic activity but not affecting the …
The switch from proteasome to immunoproteasome is increased in circulating cells of patients with fast progressive immunoglobulin A nephropathy and a…
2021
The proteasome to immunoproteasome (iPS) switch consists of β1, β2 and β5 subunit replacement by low molecular weight protein 2 (LMP2), LMP7 and multicatalytic endopeptidase-like complex-1 (MECL1) subunits, resulting in a more efficient peptide preparation for major histocompatibility complex 1 (MHC-I) presentation. It is activated by toll-like receptor (TLR) agonists and interferons and may also be influenced by genetic variation. In a previous study we found an iPS upregulation in peripheral cells of patients with immunoglobulin A nephropathy (IgAN). We aimed to investigate in 157 IgAN patients enrolled through the multinational Validation Study of the Oxford Classification of IgAN (VAL…
Biochemical Properties of Human D-Amino Acid Oxidase
2017
D-amino acid oxidase catalyzes the oxidative deamination of D-amino acids. In the brain, the NMDA receptor coagonist D-serine has been proposed as its physiological substrate. In order to shed light on the mechanisms regulating D-serine concentration at the cellular level, we biochemically characterized human DAAO (hDAAO) in greater depth. In addition to clarify the physical-chemical properties of the enzyme, we demonstrated that divalent ions and nucleotides do not affect flavoenzyme function. Moreover, the definition of hDAAO substrate specificity demonstrated that D-cysteine is the best substrate, which made it possible to propose it as a putative physiological substrate in selected tiss…