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Enzyme and Microbial Technology

Babbal, Adivitiya, Shilpa Mohanty, Yogender Pal Khasa
The exploitation of SUMO (small ubiquitin-related modifier) fusion technology at a large scale for the production of therapeutic proteins with an authentic N-terminus is majorly limited due to the higher cost of ScUlp1 protease. Therefore, the cost-effective production of Saccharomyces cerevisiae Ulp1 protease catalytic domain (402-621 aa) was targeted via its cloning under strong T7 promoter with and without histidine tag. The optimization of cultivation conditions at shake flask resulted in ScUlp1 expression of 195 mg/L in TB medium with a specific product yield of 98 mg/g DCW...
January 2019: Enzyme and Microbial Technology
Tong-Tong Tan, Xu-Dong Zhang, Zhen Miao, Ying Yu, Si-Ling Du, Xiao-Yue Hou, Jun Cai
Bacillus thuringiensis BMB181 (Bt BMB181), with high melanin production, is an ideal candidate for industrial scale production of light-stable insecticides. However, its melanogenic pathways remain unclear. In the present study, we demonstrated that Bt BMB181 failed to produce melanin after treatment with mesotrione, an inhibitor of 4-hydroxyphenylpyruvate dioxygenase in the homogentisic acid pathway of melanin synthesis. Heterologous expression experiments suggested that homogentisate-1,2-dioxygenase (HmgA) in Bt BMB171 functions normally, yet HmgA in Bt BMB181 had lost its activity, at least partly...
January 2019: Enzyme and Microbial Technology
Chan-Su Rha, Jung-Min Choi, Young Sung Jung, Eun-Ryoung Kim, Min Ji Ko, Dong-Ho Seo, Dae-Ok Kim, Cheon-Seok Park
Isoquercitrin (IQ, quercetin-3-O-β-d-glucopyranoside) has diverse biological functions, such as anti-oxidant and anti-cancer activity, but its use is limited by poor solubility and bioavailability. Enzymatically modified IQ (EMIQ) is a mixture of transglycosylated IQs that have better solubility and bioavailability than do quercetin and IQ. Two different enzymes, cyclodextrin glucanotransferase (CGTase) and amylosucrase (ASase), have the transglycosylation activity to produce EMIQ. Both enzymes produce a variety of EMIQs including IQ, IQ-glucoside (IQ-G1), IQ-diglucoside (IQ-G2), and IQ-triglucoside (IQ-G3)...
January 2019: Enzyme and Microbial Technology
Sathyanarayanan S Veeravalli, Alexander P Mathews
A novel low pH fermentation process was studied for the conversion of lactose using Lactobacillus plantarum and Lactobacillus buchneri under anoxic conditions in single co-culture, and two-stage sequential fermentations. This is aimed at producing acetate and propylene glycol (PG) as environmentally benign substitutes for currently used road and aircraft deicing chemicals. The results indicate that in the case of two-stage fermentation with immobilized L. buchneri in the second stage, lactose degradation rate increased markedly producing acetate and PG concentrations of 12...
January 2019: Enzyme and Microbial Technology
Armand Roucher, Elodie Roussarie, Régis M Gauvin, Jad Rouhana, Sébastien Gounel, Claire Stines-Chaumeil, Nicolas Mano, Rénal Backov
We present a new heterogeneous biocatalyst based on the grafting of Bilirubin Oxidase from Bacillus pumilus into macrocellular Si(HIPE) materials dedicated to water treatment. Due to the host intrinsic high porosity and monolithic character, on-line catalytic process is reached. We thus used this biocatalyst toward uni-axial flux decolorizations of Congo Red and Remazol Brilliant Blue (RBBR) at two different pH (4 and 8.2), both in presence or absence of redox mediator. In absence of redox mediators, 40% decolorization efficiency was reached within 24 h at pH 4 for Congo Red and 80% for RBBR at pH 8...
January 2019: Enzyme and Microbial Technology
Johannes Nolte, Lara-Alina Pöttgen, Julia Sperlich, Alessandra Grossert, Alexander Kempa, Nicole Teusch, Ulrich Schörken
Nordihydroguaiaretic acid (NDGA) is the major lignan of the creosote bush Larrea tridentata known for its antioxidative and pharmacological properties. Here we present the identification of glucansucrases for NDGA glucosylation and the physicochemical and biological characterization of the glucosides. Extracellular glucansucrase of L. pseudomesenteroides DSM 20193 was selected from 19 glucansucrase positive Leuconostoc and Weissella strains. Kinetic analysis of the PEG-fractionated enzyme revealed a KM of 6...
January 2019: Enzyme and Microbial Technology
Ulrike Brandt, Gulsina Galant, Christina Meinert-Berning, Alexander Steinbüchel
Thiol dioxygenases are non-heme mononuclear-iron proteins and belong to the cupin superfamily. In 2014, mercaptosuccinate dioxygenase (Msdo) of Variovorax paradoxus B4 was identified as another bacterial cysteine dioxygenase (Cdo) homolog catalyzing the conversion of mercaptosuccinate (MS) into succinate and sulfite. To gain further insights into potentially important amino acid residues for enzyme activity, seven enzyme variants were generated and analyzed. (i) Three variants comprised the substitution of one conserved histidine residue each by leucine, either supposed to be mandatory for coordination of the Fe(II) cofactor (H93 and H95) or to be important for substrate positioning within the active site (H163)...
January 2019: Enzyme and Microbial Technology
Geon-Hee Kim, Hyunwoo Jeon, Taresh P Khobragade, Mahesh D Patil, Sihyong Sung, Sanghan Yoon, Yumi Won, In Suk Choi, Hyungdon Yun
Enantiopure β-amino acids are essential precursors of various pharmaceuticals, agrochemicals and other industrially important chemicals. In this study, we selected sixteen potential ω-Transaminases (ω-TAs) by BLAST and phylogenetic tree analysis. These ω-TAs were cloned, purified and tested for their reactivity for the synthesis of model β-amino acid (R)-3-amino-4-(2,4,5-triflurophenyl) butanoic acid [3-ATfBA], a key precursor for sitagliptin. In an enzymatic cascade, lipase converted β-ketoester substrate to β-keto acid, which was subsequently aminated by the selected ω-TA to its corresponding β-amino acid...
January 2019: Enzyme and Microbial Technology
Babar K Khan, Luca Fortunato, TorOve Leiknes
Membrane-based filtration technologies have seen rapid inclusion in a variety of industrial processes, especially production of drinking water by desalination. Biological fouling of membranes is a challenge that leads to increased costs from efficiency reductions, membrane damage and ultimately, membrane replacement over time. Such costs can be mitigated by monitoring and optimizing cleaning processes for better prognosis. Monitoring bacterial accumulation in situ can therefore advance understanding of cleaning efficiency...
January 2019: Enzyme and Microbial Technology
Li Xu, Rou Wang, Hanyu Cao, Ting Xu, Lulu Han, Chundong Huang, Lingyun Jia
A facile and economical method was established for the oriented immobilization of biotin ligase (BirA) on Co3+ -NTA sepharose through H2 O2 oxidation of Co2+ and His-tag. His-tag of the BirA were designed at both N-terminal (His-BirA) and C-terminal (BirA-His), respectively. Immobilization of the His-BirA was performed, realized to 92.85% using by 10 mM H2 O2 without compromising catalytic activity. Because amounts of ions on matrix were far more than that of the immobilized BirA, EDTA should be used to remove residual ions before catalyzing, while it should be limited to lower than 30 mM, and imidazole ranging from 50 to 250 mM could be added in the catalytic system...
January 2019: Enzyme and Microbial Technology
Gabriela Leila Berto, Josman Velasco, Caio Tasso Cabos Ribeiro, Letícia Maria Zanphorlin, Mariane Noronha Domingues, Mario Tyago Murakami, Igor Polikarpov, Leandro Cristante de Oliveira, André Ferraz, Fernando Segato
Lignocellulosic materials are abundant, renewable and are emerging as valuable substrates for many industrial applications such as the production of second-generation biofuels, green chemicals and pharmaceuticals. However, the recalcitrance and the complexity of cell wall polysaccharides require multiple enzymes for their complete conversion to oligo- and monosaccharides. The endoglucanases from GH45 family are a small and relatively poorly studied group of enzymes with potential industrial application. The present study reports cloning, heterologous expression and functional characterization of two GH45 endoglucanases from mesophilic fungi Gloeophyllum trabeum (GtGH45) and thermophilic fungi Myceliophthora thermophila (MtGH45), which belong to subfamilies GH45C and GH45A, respectively...
January 2019: Enzyme and Microbial Technology
Antonielle Vieira Monclaro, Guilherme Lima Recalde, Francides Gomes da Silva, Sonia Maria de Freitas, Edivaldo Ximenes Ferreira Filho
A 22 kDa xylanase (AtXyl1) from Aspergillus tamarii was purified by two chromatographic steps and presented preference for oat spelt (OSX), birchwood (BrX) and beechwood (BeX) xylans respectively, as substrates. AtXyl1 displays the highest activity at pH 5.5 and 55 °C and showed tolerance over a range of different phenolic compounds. The activity of AtXyl1 was not inhibited when the enzyme was incubated with ferulic acid (FA) using OSX or BrX as substrate. On the other hand, the incubation of AtXyl1 with BeX and FA resulted in an increase in enzyme activity...
January 2019: Enzyme and Microbial Technology
Juan Han, Yunfeng Cai, Yun Wang, Lei Gu, Chunmei Li, Yanli Mao, Wenli Zhang, Liang Ni
In this work, a novel Ni2+ &AABOB bifunctional core-shell magnetic composite nanoparticles (MNPs), Fe3 O4 @p(AABOB-co-AIM-Ni2+ ), were synthesized for the immobilization of horseradish peroxidase (HRP). Two kinds of interaction are simultaneously incorporated to improve the stability and reusability of immobilized HRP: one is the coordination between Ni2+ and amide group in HRP, and the other is the interaction of the wulff-type boronic acid group with cis-diol-containing HRP which belongs to a type of glycoproteins...
January 2019: Enzyme and Microbial Technology
Io Antonopoulou, Laura Iancu, Peter Jütten, Alexander Piechot, Ulrika Rova, Paul Christakopoulos
The feruloyl esterases Fae125, Fae7262 and Fae68 from Talaromyces wortmannii were screened in 10 different solvent: buffer systems in terms of residual hydrolytic activity and of the ability for the transesterification of vinyl ferulate with prenol or l-arabinose. Among the tested enzymes, the acetyl xylan-related Fae125 belonging to the phylogenetic subfamily 5 showed highest yield and selectivity for both products in alkane: buffer systems (n-hexane or n-octane). Response surface methodology, based on a 5-level and 6-factor central composite design, revealed that the substrate molar ratio and the water content were the most significant variables for the bioconversion yield and selectivity...
January 2019: Enzyme and Microbial Technology
Hafezeh Salehabadi, Khosro Khajeh, Bahareh Dabirmanesh, Mahmood Biglar, Massoud Amanlou
Surface plasmon resonance (SPR) biosensor has been utilized for monitoring analyte-ligand interactions in modern drug discovery processes. SPR biosensors measure the change in refractive indexes over the course of analyte molecules' binding to a specific immobilized ligand on sensor chip. This effort highlights a comprehensive SPR study besides enzymatic assay for discovery of new Angiotensin Converting Enzyme (ACE) inhibitors via screening of medicinal plants. At first, five medicinal plants were selected as potential sources for developing new ACE inhibitors through hydrolyzing hippuryl-L-histidyl-L-leucine (HHL) assay...
January 2019: Enzyme and Microbial Technology
Péter Polyák, Eszter Urbán, Gergely N Nagy, Beáta G Vértessy, Béla Pukánszky
The enzyme catalyzed degradation of poly(3-hydroxybutyrate) (PHB) is a two-step process consisting of the adsorption of the enzyme on the surface of a PHB substrate and the cleavage of ester bonds. A deactivated enzyme was prepared by point mutagenesis to separate the two steps from each other. Measurements carried out with active and inactive enzymes on PHB particles proved that mutagenesis was successful and the modified enzyme did not catalyze degradation. Based on the Michaelis-Menten approach a kinetic model was proposed which could describe the processes quantitatively, the agreement between prediction and the measured data was excellent...
January 2019: Enzyme and Microbial Technology
Shanmugasundaram Senthil Balan, Panagal Mani, C Ganesh Kumar, Singaram Jayalakshmi
Marine microbes have gained significant attention as potential biofactories for broad spectrum bioactive compounds. In the recent years, bioactive biosurfactants have warranted renewed interest from both environmental and clinical sectors as anti-biofilm agents due to their excellent properties of dispersing microbial biofilms. The present study explores a new glycolipid biosurfactant produced by a marine Staphylococcus saprophyticus exhibiting interesting biological activities. This glycolipid biosurfactant was purified and identified as Mannose-Mannose-Oleic acid (named as Staphylosan) based on the results of NMR, GC, GC-MS, MALDI-TOF-MS and tandem MS analysis...
January 2019: Enzyme and Microbial Technology
Gang Xu, Qiang Zhao, Bin Huang, Jinghui Zhou, Fuxiang Cao
Penicillin acylase is commonly used to produce the medical intermediates of 6-Aminopenicillanic acid (6-APA) and 7-Aminodesacetoxycephalosporanic acid (7-ADCA) in industrial process. Nowadays, Penicillin G acylase (PGA) has been widely applied for making pharmaceutical intermediates, while penicillin V acylase (PVA) has been less used for that due to its low activity and poor conversion. In this study, a PVA from Bacillus sphaericus (BspPVA) was employed for directed evolution study with hoping to increase its catalytic efficiency...
December 2018: Enzyme and Microbial Technology
Fan Li, Yuanyuan Yu, Qiang Wang, Jiugang Yuan, Ping Wang, Xuerong Fan
Polydopamine (PDA) films are generally produced by spontaneous oxidation of dopamine in basic medium, which is a highly convenient method for surface modification. This method is, however, not suitable for alkaline sensitive materials, which have poor stability in alkali medium and need over-long reaction time. Herein, we compared two dopamine polymerization methods: conventional method (induced by O2 at pH 8.5) and laccase-catalyzed method (at pH 5.5). The comparison showed that the polymerization of dopamine was greatly improved when catalyzed by laccase...
December 2018: Enzyme and Microbial Technology
Xiuwen Wang, Aixia Wang, Lingfeng Zhu, Dongliang Hua, Jiayang Qin
A sufficient supply of reducing equivalents is essential for obtaining the maximum yield of target products in anaerobic fermentation. The pyruvate dehydrogenase (PDH) complex controls the critical step in pyruvate conversion to acetyl-CoA and NADH. However, in anaerobic Escherichia coli, PDH residing in the dihydrolipoamide dehydrogenase (LPD) component is normally inactive due to inhibition by NADH. In this study, the protein engineering of LPD by structural analysis was explored to eliminate this inhibition...
December 2018: Enzyme and Microbial Technology
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