scholarly journals Exploring the Diversity of Fungal DyPs in Mangrove Soils to Produce and Characterize Novel Biocatalysts

2021 ◽  
Vol 7 (5) ◽  
pp. 321
Author(s):  
Amal Ben Ayed ◽  
Geoffroy Saint-Genis ◽  
Laurent Vallon ◽  
Dolores Linde ◽  
Annick Turbé-Doan ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
Chemical Properties ◽  
Avicennia Marina ◽  
Reactive Black 5 ◽  
Wet Season ◽  
Rhizophora Stylosa ◽  
Genes Encoding ◽  
Wide Ph Range ◽  
Ph Range

The functional diversity of the New Caledonian mangrove sediments was examined, observing the distribution of fungal dye-decolorizing peroxidases (DyPs), together with the complete biochemical characterization of the main DyP. Using a functional metabarcoding approach, the diversity of expressed genes encoding fungal DyPs was investigated in surface and deeper sediments, collected beneath either Avicennia marina or Rhizophora stylosa trees, during either the wet or the dry seasons. The highest DyP diversity was observed in surface sediments beneath the R. stylosa area during the wet season, and one particular operational functional unit (OFU1) was detected as the most abundant DyP isoform. This OFU was found in all sediment samples, representing 51–100% of the total DyP-encoding sequences in 70% of the samples. The complete cDNA sequence corresponding to this abundant DyP (OFU 1) was retrieved by gene capture, cloned, and heterologously expressed in Pichia pastoris. The recombinant enzyme, called DyP1, was purified and characterized, leading to the description of its physical–chemical properties, its ability to oxidize diverse phenolic substrates, and its potential to decolorize textile dyes; DyP1 was more active at low pH, though moderately stable over a wide pH range. The enzyme was very stable at temperatures up to 50 °C, retaining 60% activity after 180 min incubation. Its ability to decolorize industrial dyes was also tested on Reactive Blue 19, Acid Black, Disperse Blue 79, and Reactive Black 5. The effect of hydrogen peroxide and sea salt on DyP1 activity was studied and compared to what is reported for previously characterized enzymes from terrestrial and marine-derived fungi.

scholarly journals Biochemical Characterization of a New β-Agarase from Cellulophaga Algicola

2019 ◽  
Vol 20 (9) ◽  
pp. 2143 ◽  
Author(s):  
Han ◽  
Zhang ◽  
Yang
Keyword(s):  
Structural Model ◽  
Biochemical Characterization ◽  
Maximum Activity ◽  
Optimal Temperature ◽  
Salt Tolerant ◽  
Wide Ph Range ◽  
Specific Activities ◽  
Ph Range ◽  
Layer Chromatography

Cellulophaga algicola DSM 14237, isolated from the Eastern Antarctic coastal zone, was found to be able to hydrolyze several types of polysaccharide materials. In this study, a predicted β-agarase (CaAga1) from C. algicola was heterologously expressed in Escherichia coli. The purified recombinant CaAga1 showed specific activities of 29.39, 20.20, 14.12, and 8.99 U/mg toward agarose, pure agar, and crude agars from Gracilaria lemaneiformis and Porphyra haitanensis, respectively. CaAga1 exhibited an optimal temperature and pH of 40 oC and 7, respectively. CaAga1 was stable over a wide pH range from 4 to 11. The recombinant enzyme showed an unusual thermostability, that is, it was stable at temperature below or equal to 40oC and around 70 oC, but was thermolabile at about 50 oC. With the agarose as the substrate, the Km and Vmax values for CaAga1 were 1.19 mg/mL and 36.21 U/mg, respectively. The reducing reagent (dithiothreitol) enhanced the activity of CaAga1 by more than one fold. In addition, CaAga1 was salt-tolerant given that it retained approximately 70% of the maximum activity in the presence of 2 M NaCl. The thin layer chromatography results indicated that CaAga1 is an endo-type β-agarase and efficiently hydrolyzed agarose into neoagarotetraose (NA4) and neoagarohexaose (NA6). A structural model of CaAga1 in complex with neoagarooctaose (NA8) was built by homology modeling and explained the hydrolysis pattern of CaAga1.

Biochemical and functional characterization of OsCSD3, a novel CuZn superoxide dismutase from rice

2018 ◽  
Vol 475 (19) ◽  
pp. 3105-3121 ◽  
Author(s):  
Ravi Prakash Sanyal ◽  
Amol Samant ◽  
Vishal Prashar ◽  
Hari Sharan Misra ◽  
Ajay Saini
Keyword(s):  
Oxidative Stress ◽  
Thermal Inactivation ◽  
Biochemical Characterization ◽  
Spectral Characteristics ◽  
Functional Characterization ◽  
Double Knockout ◽  
Wide Ph Range ◽  
Ph Range

Superoxide dismutases (SODs, EC 1.15.1.1) belong to an important group of antioxidant metalloenzymes. Multiple SODs exist for scavenging of reactive oxygen species (ROS) in different cellular compartments to maintain an intricate ROS balance. The present study deals with molecular and biochemical characterization of CuZn SOD encoded by LOC_Os03g11960 (referred to as OsCSD3), which is the least studied among the four rice isozymes. The OsCSD3 showed higher similarity to peroxisomal SODs in plants. The OsCSD3 transcript was up-regulated in response to salinity, drought, and oxidative stress. Full-length cDNA encoding OsCSD3 was cloned and expressed in Escherichia coli and analyzed for spectral characteristics. UV (ultraviolet)–visible spectroscopic analysis showed evidences of d–d transitions, while circular dichroism analysis indicated high β-sheet content in the protein. The OsCSD3 existed as homodimer (∼36 kDa) with both Cu2+ and Zn2+ metal cofactors and was substantially active over a wide pH range (7.0–10.8), with optimum pH of 9.0. The enzyme was sensitive to diethyldithiocarbamate but insensitive to sodium azide, which are the characteristics features of CuZn SODs. The enzyme also exhibited bicarbonate-dependent peroxidase activity. Unlike several other known CuZn SODs, OsCSD3 showed higher tolerance to hydrogen peroxide and thermal inactivation. Heterologous overexpression of OsCSD3 enhanced tolerance of E. coli sod double-knockout (ΔsodA ΔsodB) mutant and wild-type strain against methyl viologen-induced oxidative stress, indicating the in vivo function of this enzyme. The results show that the locus LOC_Os03g11960 of rice encodes a functional CuZn SOD with biochemical characteristics similar to the peroxisomal isozymes.

Biochemical characterization of a glucoamylase from Saccharomycopsis fibuligera R64

Biologia ◽  
2011 ◽  
Vol 66 (1) ◽  
Author(s):  
Dessy Natalia ◽  
Keni Vidilaseris ◽  
Pasjan Satrimafitrah ◽  
Wangsa Ismaya ◽  
Purkan ◽  
...  
Keyword(s):  
Mass Spectrometry ◽  
Amino Acid ◽  
Amino Acid Sequence Identity ◽  
Biochemical Characterization ◽  
Saccharomycopsis Fibuligera ◽  
Sequence Identity ◽  
Ic50 Value ◽  
High Amino Acid ◽  
Ph Range

AbstractGlucoamylase from the yeast Saccharomycopsis fibuligera R64 (GLL1) has successfully been purified and characterized. The molecular mass of the enzyme was 56,583 Da as determined by mass spectrometry. The purified enzyme demonstrated optimum activity in the pH range of 5.6–6.4 and at 50°C. The activity of the enzyme was inhibited by acarbose with the IC50 value of 5 μM. GLL1 shares high amino acid sequence identity with GLU1 and GLA1, which are Saccharomycopsis fibuligera glucoamylases from the strains HUT7212 and KZ, respectively. The properties of GLL1, however, resemble that of GLU1. The elucidation of the primary structure of GLL1 contributes to the explanation of this finding.

scholarly journals Identification and biochemical characterization of a novel PP2C-like Ser/Thr phosphatase inE. coli

2018 ◽  
Author(s):  
Krithika Rajagopalan ◽  
Jonathan Dworkin
Keyword(s):  
Biochemical Characterization ◽  
Regulatory Protein ◽  
Biochemical Properties ◽  
Bacterial Genomes ◽  
Phosphatase Inhibitors ◽  
E Coli ◽  
Specificity And Sensitivity ◽  
Genes Encoding ◽  
Number Of Bacteria

AbstractIn bacteria, signaling phosphorylation is thought to occur primarily on His and Asp residues. However, phosphoproteomic surveys in phylogenetically diverse bacteria over the past decade have identified numerous proteins that are phosphorylated on Ser and/or Thr residues. Consistently, genes encoding Ser/Thr kinases are present in many bacterial genomes such asE. coli,which encodes at least three Ser/Thr kinases. Since Ser/Thr phosphorylation is a stable modification, a dedicated phosphatase is necessary to allow reversible regulation. Ser/Thr phosphatases belonging to several conserved families are found in bacteria. One family of particular interest are Ser/Thr phosphatases which have extensive sequence and structural homology to eukaryotic Ser/Thr PP2C phosphatases. These proteins, called eSTPs (eukaryotic-like Ser/Thr phosphatases), have been identified in a number of bacteria, but not inE. coli.Here, we describe a previously unknown eSTP encoded by anE. coliORF,yegK,and characterize its biochemical properties including its kinetics, substrate specificity and sensitivity to known phosphatase inhibitors. We investigate differences in the activity of this protein in closely relatedE. colistrains. Finally, we demonstrate that this eSTP acts to dephosphorylate a novel Ser/Thr kinase which is encoded in the same operon.ImportanceRegulatory protein phosphorylation is a conserved mechanism of signaling in all biological systems. Recent phosphoproteomic analyses of phylogenetically diverse bacteria including the model Gram-negative bacteriumE. colidemonstrate that many proteins are phosphorylated on serine or threonine residues. In contrast to phosphorylation on histidine or aspartate residues, phosphorylation of serine and threonine residues is stable and requires the action of a partner Ser/Thr phosphatase to remove the modification. Although a number of Ser/Thr kinases have been reported inE. coli, no partner Ser/Thrphosphatases have been identified. Here, we biochemically characterize a novel Ser/Thr phosphatase that acts to dephosphorylate a Ser/Thr kinase that is encoded in the same operon.

scholarly journals Biochemical characterization of the tetrahydrobiopterin synthesis pathway in the oleaginous fungus Mortierella alpina

Microbiology ◽  
2011 ◽  
Vol 157 (11) ◽  
pp. 3059-3070 ◽  
Author(s):  
Hongchao Wang ◽  
Bo Yang ◽  
Guangfei Hao ◽  
Yun Feng ◽  
Haiqin Chen ◽  
...  
Keyword(s):  
Biochemical Characterization ◽  
De Novo ◽  
Mortierella Alpina ◽  
Recombinant Enzymes ◽  
Homologous Proteins ◽  
Genes Encoding ◽  
Oleaginous Fungus ◽  
Cell Processes ◽  
Common Substrate

We characterized the de novo biosynthetic pathway of tetrahydrobiopterin (BH4) in the lipid-producing fungus Mortierella alpina. The BH4 cofactor is essential for various cell processes, and is probably present in every cell or tissue of higher organisms. Genes encoding two copies of GTP cyclohydrolase I (GTPCH-1 and GTPCH-2) for the conversion of GTP to dihydroneopterin triphosphate (H2-NTP), 6-pyruvoyltetrahydropterin synthase (PTPS) for the conversion of H2-NTP to 6-pyruvoyltetrahydropterin (PPH4), and sepiapterin reductase (SR) for the conversion of PPH4 to BH4, were expressed heterologously in Escherichia coli. The recombinant enzymes were produced as His-tagged fusion proteins and were purified to homogeneity to investigate their enzymic activities. Enzyme products were analysed by HPLC and electrospray ionization-MS. Kinetic parameters and other properties of GTPCH, PTPS and SR were investigated. Physiological roles of BH4 in M. alpina are discussed, and comparative analyses between GTPCH, PTPS and SR proteins and other homologous proteins were performed. The presence of two functional GTPCH enzymes has, as far as we are aware, not been reported previously, reflecting the unique ability of this fungus to synthesize both BH4 and folate, using the GTPCH product as a common substrate. To our knowledge, this study is the first to report the comprehensive characterization of a BH4 biosynthesis pathway in a fungus.

scholarly journals Characterization of the bacteriocin-producing strain Lactobacillus paracasei subsp. paracasei BGUB9

2010 ◽  
Vol 62 (4) ◽  
pp. 889-899 ◽  
Author(s):  
Maja Tolinacki ◽  
M. Kojic ◽  
Jelena Lozo ◽  
Amarela Terzic-Vidojevic ◽  
L. Topisirovic ◽  
...  
Keyword(s):  
Antimicrobial Activity ◽  
Biochemical Characterization ◽  
Proteolytic Enzymes ◽  
Lactobacillus Paracasei ◽  
Plasmid Curing ◽  
Closely Related Species ◽  
Antimicrobial Spectrum ◽  
Activity Test ◽  
Ph Range

The strain Lactobacillus paracasei subsp. paracasei BGUB9 that was isolated from traditionally homemade hard cheese produces bacteriocin designated as BacUB9, with an approximate molecular mass of 4 kDa. Biochemical characterization and the antimicrobial activity test of BacUB9 were performed. The onset of BacUB9 biosynthesis was observed at the end of an exponential phase of growth. Bacteriocin UB9 retained the antimicrobial activity within the pH range from 1 to 10 and after treatment at 100oC for 30 min. The bacteriocin is susceptible to the activity of proteolytic enzymes. Bacteriocin BacUB9 has a very narrow antimicrobial spectrum, limited to several strains that belong to closely related species. The effect of BGUB9 on the growth of the strain Lactobacillus paracasei subsp. paracasei BGHN14 in a mixed culture was monitored. The mode of action of BacUB9 on the strain BGHN14 was identified as bacteriostatic. Plasmid curing results indicated that a plasmid, designated as pUB9, seemed to be responsible for both bacteriocin BacUB9 production and host immunity.

scholarly journals Withdrawal: Functional and biochemical characterization of cucumber genes encoding two copper ATPases CsHMA5.1 and CsHMA5.2.

2019 ◽  
Vol 294 (35) ◽  
pp. 13200-13200
Author(s):  
Magdalena Migocka ◽  
Ewelina Posyniak ◽  
Ewa Maciaszczyk-Dziubinska ◽  
Anna Papierniak ◽  
Anna Kosieradzaka
Keyword(s):  
Biochemical Characterization ◽  
Copper Atpases ◽  
Genes Encoding

GENOME-WIDE IDENTIFICATION AND COMPUTATIONAL CHARACTERIZATION OF THE NUCLEAR FACTOR-YC SUB-UNITS IN GRAIN AMARANTH (Amaranthus hypochondriacus)

2021 ◽  
Vol 66 (3) ◽  
pp. 161-169
Author(s):  
Huyen Tran Thi Thanh ◽  
Hong La Viet ◽  
Quynh Le Thi Ngoc ◽  
Thuy Pham Chau ◽  
Quyen Ha Thi ◽  
...  
Keyword(s):  
Growth And Development ◽  
Expression Profiles ◽  
Functional Characterization ◽  
Chemical Properties ◽  
Nuclear Factor ◽  
Grain Amaranth ◽  
Amaranthus Hypochondriacus ◽  
Grand Average ◽  
Genes Encoding

Nuclear factor-Y (NF-Y) has been known as one of the plant-specific transcription factors that play key roles in numerous biological processes during the growth and development of plant species. In this study, a comprehensive analysis of NF-YC sub-units in grain amaranth (Amaranthus hypochondriacus) was carried out based on the bioinformatics approaches. Firstly, a total of five members of the NF-YC sub-units was reported in the grain amaranth. Its structural analyses revealed that the NF-YC sub-units were variable in physic-chemical properties, like protein sizes, molecular masses, isoelectric point, instability index, and grand average of hydropathy. Of our interest, the expression profiles of genes encoding NF-YC sub-units in various tissues\organs during the growth and development of grain amaranth. We found that three genes, including AhNF-YC01, AhNF-YC04, and AhNF-YC05 were highly expressed in leaf, root, floral, immature seed, and stem tissues. Interestingly, AhNF-YC05 was exclusively expressed in leaf and stem tissues. Taken together, our study could provide a solid understanding for further functional characterization of genes encoding NF-YC sub-units in grain amaranth.

scholarly journals A Minimally Invasive Microsensor Specially Designed for Simultaneous Dissolved Oxygen and pH Biofilm Profiling

Sensors ◽  
2019 ◽  
Vol 19 (21) ◽  
pp. 4747 ◽  
Author(s):  
Xavier Guimerà ◽  
Ana Moya ◽  
Antonio David Dorado ◽  
Xavi Illa ◽  
Rosa Villa ◽  
...  
Keyword(s):  
Minimally Invasive ◽  
Dissolved Oxygen ◽  
Ph Monitoring ◽  
Ph Sensing ◽  
Wide Ph Range ◽  
Ph Range ◽  
Biofilm Structure ◽  
Parallel Arrays ◽  
Ph Electrodes

A novel sensing device for simultaneous dissolved oxygen (DO) and pH monitoring specially designed for biofilm profiling is presented in this work. This device enabled the recording of instantaneous DO and pH dynamic profiles within biofilms, improving the tools available for the study and the characterization of biological systems. The microsensor consisted of two parallel arrays of microelectrodes. Microelectrodes used for DO sensing were bare gold electrodes, while microelectrodes used for pH sensing were platinum-based electrodes modified using electrodeposited iridium oxide. The device was fabricated with a polyimide (Kapton®) film of 127 µm as a substrate for minimizing the damage caused on the biofilm structure during its insertion. The electrodes were covered with a Nafion® layer to increase sensor stability and repeatability and to avoid electrode surface fouling. DO microelectrodes showed a linear response in the range 0–8 mg L−1, a detection limit of 0.05 mg L−1, and a sensitivity of 2.06 nA L mg−1. pH electrodes showed a linear super-Nernstian response (74.2 ± 0.7 mV/pH unit) in a wide pH range (pH 4−9). The multi-analyte sensor array was validated in a flat plate bioreactor where simultaneous and instantaneous pH and DO profiles within a sulfide oxidizing biofilm were recorded. The electrodes spatial resolution, the monitoring sensitivity, and the minimally invasive features exhibited by the proposed microsensor improved biofilm monitoring performance, enabling the quantification of mass transfer resistances and the assessment of biological activity.

Effect of speciation transformation on the coagulation behavior of Al13 and Al13 aggregates

2009 ◽  
Vol 59 (4) ◽  
pp. 815-822 ◽  
Author(s):  
Xiaohong Wu ◽  
Changqing Ye ◽  
Dongsheng Wang ◽  
Xiaopeng Ge ◽  
Hongxiao Tang
Keyword(s):  
Dominant Species ◽  
Chemical Interaction ◽  
Surface Adsorption ◽  
Surface Precipitation ◽  
Floc Size ◽  
Species Transformation ◽  
Wide Ph Range ◽  
Ph Range

Flocculation of kaolin suspension with aluminium fractal polycations was investigated as a function of aluminium concentration and pH. Aluminium flocculants included Al13 and Al13 aggregates with OH/Al ratio of 2.6 and 2.8, respectively. The flocculation kinetics and floc size distribution were monitored by light scattering. The characterization of flocculants showed that the tridecatmer Al13 and bridged [Al13]n with out-sphere structure were the dominant species for all flocculants in a wide pH range. The coagulation results indicated that the pre and in situ-formed [Al13]n play a key role in removing particles. With the increasing concentration of [Al13]n, coagulation mechanisms were transformed from charge-neutralization, electro-patch coagulation to bridge-aggregation. Moreover, sweep-flocculation was involved at higher dosage besides other three mechanisms when amorphous aluminium oxides formed. Hence, chemical interaction between particles and flocculants evolved from surface adsorption to surface precipitation for aluminium polycations by virtue of species transformation.

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