scholarly journals Improvement of the Transglycosylation Efficiency of a Lacto-N-Biosidase from Bifidobacterium bifidum by Protein Engineering

2021 ◽  
Vol 11 (23) ◽  
pp. 11493
Author(s):  
Marlene Vuillemin ◽  
Jesper Holck ◽  
Martin Matwiejuk ◽  
Eduardo S. Moreno Prieto ◽  
Jan Muschiol ◽  
...  
Keyword(s):  
Human Milk ◽  
Target Position ◽  
Product Formation ◽  
Bifidobacterium Bifidum ◽  
Donor Substrate ◽  
Transglycosylation Activity ◽  
Initial Substrate ◽  
Major Reaction Product ◽  
Major Reaction ◽  
Substrate Concentrations

The lacto-N-biosidase LnbB from Bifidobacterium bifidum JCM 1254 was engineered to improve its negligible transglycosylation efficiency with the purpose of enzymatically synthesizing lacto-N-tetraose (LNT; Gal-β1,3-GlcNAc-β1,3-Gal-β1,4-Glc) in one enzymatic step. LNT is a prebiotic human milk oligosaccharide in itself and constitutes the structural core of a range of more complex human milk oligosaccharides as well. Thirteen different LnbB variants were expressed and screened for transglycosylation activity by monitoring transglycosylation product formation using lacto-N-biose 1,2-oxazoline as donor substrate and lactose as acceptor substrate. LNT was the major reaction product, yet careful reaction analysis revealed the formation of three additional LNT isomers, which we identified to have a β1,2-linkage, a β1,6-linkage, and a 1,1-linkage, respectively, between lacto-N-biose (Gal-β1,3-GlcNAc) and lactose. Considering both maximal transglycosylation yield and regioselectivity as well as minimal product hydrolysis, the best variant was LnbB W394H, closely followed by W465H and Y419N. A high transglycosylation yield was also obtained with W394F, yet the substitution of W394 and W465 of the subsite −1 hydrophobic platform in the enzyme with His dramatically impaired the undesirable product hydrolysis as compared to substitution with Phe; the effect was most pronounced for W465. Using p-nitrophenyl-β-lacto-N-bioside as donor substrate manifested W394 as an important target position. The optimization of the substrate concentrations confirmed that high initial substrate concentration and high acceptor-to-donor ratio both favor transglycosylation.

THE INFLUENCE OF ETHANOL ON THE CONVERSION OF PREGNENOLONE TO PROGESTERONE BY HUMAN PLACENTAL MICROSOMES

1974 ◽  
Vol 76 (1) ◽  
pp. 178-188 ◽  
Author(s):  
H. Lübbert ◽  
K. Pollow ◽  
R. Wagner ◽  
J. Hammerstein
Keyword(s):  
Kinetic Parameters ◽  
Enzyme Preparation ◽  
Ethanol Concentration ◽  
Hydroxysteroid Dehydrogenase ◽  
Product Formation ◽  
Linear Increase ◽  
Maximal Velocity ◽  
Microsomal Enzyme ◽  
Temperature Optima ◽  
Substrate Concentrations

ABSTRACT The effects of ethanol on kinetic parameters of placental Δ5-3β-hydroxysteroid dehydrogenase were studied. In the presence of high pregnenolone concentrations (50 μm, [S] > Km) the microsomal enzyme preparation exhibited an almost linear increase in activity as the ethanol concentration in the medium was raised from 2.5 to 15 % (v/v). At lower substrate concentrations ([S] << Km) ethanol caused inhibition. Other effects of ethanol were: linearity of product formation with time was prolonged; the maximal velocity was markedly increased; the Km for pregnenolone slightly decreased with increasing ethanol concentrations (2.5 to 10 %, v/v) whereas the Km for NAD remained the same. The pH and temperature optima of the reaction were unaffected by ethanol. Other organic solvents caused similar effects.

Phosphorus Release Kinetics in Biofilm Reactors

1992 ◽  
Vol 26 (3-4) ◽  
pp. 567-576 ◽  
Author(s):  
F. A. Ruiz-Treviño ◽  
S. González-Martínez ◽  
C. Doria-Serrano ◽  
M. Hernández-Esparza
Keyword(s):  
Phosphorus Removal ◽  
Release Kinetics ◽  
Phosphorus Release ◽  
Organic Substrate ◽  
Biofilm Reactor ◽  
Substrate Uptake ◽  
Biofilm Reactors ◽  
Initial Substrate ◽  
Linear Behaviour ◽  
Substrate Concentrations

This paper presents the kinetic analysis, using Generalized Power-Law equations to describe the results of an experimental investigation conducted on a batch submerged biofilm reactor for phosphorus removal under an anaerobic/aerobic cycle. The observed rates and amounts of phosphorus release and organic substrate uptake in the anaerobic phase leads to a kinetic model in which these two variables are dependent on each other with a non-linear behaviour and reach equilibrium values in both cases, at different times and are function of rate constants ratio. The model has a good fit with experimental data except for C uptake at anaerobic contact times longer than four hours, where other kinetics are implied. Kinetic parameters were obtained with different initial substrate concentrations, anaerobic contact cycles, and type of substrates.

METHYL 3-NITRO- AND 3-AMINO-3-DEOXY-β-D-GALACTOPYRANOSIDE AND -MANNOPYRANOSIDE

1963 ◽  
Vol 41 (6) ◽  
pp. 1606-1611 ◽  
Author(s):  
Hans Helmut Baer ◽  
Frank Kienzle
Keyword(s):  
Crystalline State ◽  
Major Reaction Product ◽  
Major Reaction ◽  
Amino Derivatives ◽  
Facile Preparation ◽  
Hydrolysis Of

The steric course of the nitromethane cyclization of L′-methoxy-D-hydroxymethyldiglycolic aldehyde was investigated. Methyl 3-nitro-3-deoxy-β-D-galactopyranoside was shown to arise as a second major reaction product in addition to the previously isolated principal stereoisomer, the gluco derivative. The corresponding manno stereoisomer is formed to a smaller extent. The configurations of the new methyl nitrodeoxyglycosides were established by conversion into the corresponding amino derivatives and hydrolysis of these latter to the known 3-amino-3-deoxy-D-galactose and -D-mannose hydrochlorides. All the products were obtained in a crystalline state. The reaction lends itself to a facile preparation of the nitrogenous galactose derivatives.

scholarly journals Growth of bifidobacteria in mammalian milk

2013 ◽  
Vol 58 (No. 3) ◽  
pp. 99-105 ◽  
Author(s):  
Š. Ročková ◽  
V. Rada ◽  
J. Havlík ◽  
R. Švejstil ◽  
E. Vlková ◽  
...  
Keyword(s):  
Lactic Acid ◽  
Human Milk ◽  
Bacterial Species ◽  
Intestinal Bacteria ◽  
Bifidobacterium Bifidum ◽  
Microbial Colonization ◽  
Animal Origin ◽  
Good Growth ◽  
Animal Milk ◽  
Species Specific

Microbial colonization of the mammalian intestine begins at birth, when from a sterile state a newborn infant is exposed to an external environment rich in various bacterial species. An important group of intestinal bacteria comprises bifidobacteria. Bifidobacteria represent major intestinal microbiota during the breast-feeding period. Animal milk contains all crucial nutrients for babies&rsquo; intestinal microflora. The aim of our work was to test the influence of different mammalian milk on the growth of bifidobacteria. The growth of seven strains of bifidobacteria in human milk, the colostrum of swine, cow&rsquo;s milk, sheep&rsquo;s milk, and rabbit&rsquo;s milk was tested. Good growth accompanied by the production of lactic acid was observed not only in human milk, but also in the other kinds of milk in all three strains of Bifidobacterium bifidum of different origin. Human milk selectively supported the production of lactic acid of human bifidobacterial isolates, especially the Bifidobacterium bifidum species. The promotion of bifidobacteria by milk is species-specific. Human milk contains a key factor for the growth of specific species or strains of human-origin bifidobacteria compared to other kinds of milk. In contrast, some components (maybe lysozyme) of human milk inhibited the growth of Bifidobacterium animalis. Animal-origin strains of bifidobacteria were not able to significantly grow even in milk of animal origin, with the exception of B. animalis subsp. lactis&nbsp;1,2, which slightly grew in sheep&rsquo;s milk.

Hydrogen Peroxide Decay in Waters with Suspended Soils: Evidence for Biologically Mediated Processes

1990 ◽  
Vol 47 (5) ◽  
pp. 888-893 ◽  
Author(s):  
William J. Cooper ◽  
Richard G. Zepp
Keyword(s):  
Hydrogen Peroxide ◽  
Natural Water ◽  
Water Samples ◽  
First Order ◽  
Natural Water Samples ◽  
Major Reaction Product ◽  
Major Reaction ◽  
Chemical Sterilization ◽  
Pseudo First Order ◽  
Catalyzed Oxidation

Hydrogen peroxide decay studies have been conducted in suspensions of several well-characterized soils and in natural water samples. Kinetic and product studies indicated that the decay was biologically-mediated, and could be described by pseudo first-order rate expressions. At an initial H2O2 concentration of 0.5 μM, the hydrogen peroxide half-life varied from 1 to 8 h. The decay was inhibited by thermal and chemical sterilization of the soils. Peroxidase activity was inferred in several natural water samples, where the suspended particles catalyzed the oxidation of p-anisidine by hydrogen peroxide. The mass spectrum of the major reaction product indicated that it was the dimer, possibly benzoquinone-4-methoxyanil, a product that also was observed from the horseradish peroxidase-catalyzed oxidation of p-anisidine by hydrogen peroxide.

scholarly journals Production of Sugar Feedstocks for Fermentation Processes from Selected Fast Growing Grasses

Energies ◽  
2019 ◽  
Vol 12 (16) ◽  
pp. 3129 ◽  
Author(s):  
Kamila Przybysz ◽  
Edyta Małachowska ◽  
Danuta Martyniak ◽  
Piotr Boruszewski ◽  
Halina Kalinowska ◽  
...  
Keyword(s):  
Reducing Sugars ◽  
Kraft Pulping ◽  
Cellulose Content ◽  
Potential Yield ◽  
Fermentation Processes ◽  
Fast Growing ◽  
Initial Substrate ◽  
Scientific Goal ◽  
Hydrolysis Of ◽  
Substrate Concentrations

This study showed that kraft cellulosic pulps from Miscanthus giganetus JM Greef and Deuter ex Hodk. and Renvoize, sweet sorghum and 5 other fast growing grasses may be easily enzymatically converted to glucose-rich sugar feedstocks. The scientific goal of the paper was to assess and compare the potential yield of hydrolysis and verify whether these grasses may be a source of sugars for fermentation processes. Kraft pulping was used as a pretreatment method and hydrolysis of the pulps was conducted using a commercial multienzyme preparation containing cellulases and xylanases at initial substrate concentrations of 0.476, 3.88 and 7.46% w/v, and 3 different enzyme loadings. Results showed that tall wheatgrass, striped tuber oat grass, tall fescue and smooth bromegrass may be efficiently converted to sugar feedstocks for biotechnology application, but that the simple reducing sugars yield is lower than for wood, due to lower cellulose content.

scholarly journals One-pot synthesis of substituted pyrrolo[3,4-b]pyridine-4,5-diones based on the reaction of N-(1-(4-hydroxy-6-methyl-2-oxo-2H-pyran-3-yl)-2-oxo-2-arylethyl)acetamide with amines

2019 ◽  
Vol 15 ◽  
pp. 2840-2846
Author(s):  
Valeriya G Melekhina ◽  
Andrey N Komogortsev ◽  
Boris V Lichitsky ◽  
Vitaly S Mityanov ◽  
Artem N Fakhrutdinov ◽  
...  
Keyword(s):  
Starting Material ◽  
Primary Amines ◽  
Synthetic Method ◽  
One Pot ◽  
One Pot Synthesis ◽  
Major Reaction Product ◽  
Major Reaction

The condensation of primary amines with N-(1-(4-hydroxy-6-methyl-2-oxo-2H-pyran-3-yl)-2-oxo-2-arylethyl)acetamides was explored. Thus, a previously unknown recyclization of the starting material was observed in acidic ethanol in the presence of an amine, which provided the corresponding dihydropyrrolone derivative as the major reaction product. Based on this transformation, a practical and convenient one-pot synthetic method for substituted pyrrolo[3,4-b]pyridin-5-ones could be devised.

scholarly journals Human Deoxycytidine Kinase Is a Valuable Biocatalyst for the Synthesis of Nucleotide Analogues

Catalysts ◽  
2019 ◽  
Vol 9 (12) ◽  
pp. 997 ◽  
Author(s):  
Katja F. Hellendahl ◽  
Sarah Kamel ◽  
Albane Wetterwald ◽  
Peter Neubauer ◽  
Anke Wagner
Keyword(s):  
Substrate Concentration ◽  
Food Additives ◽  
Product Yield ◽  
Building Blocks ◽  
Product Formation ◽  
Industrial Applications ◽  
Modified Nucleosides ◽  
Deoxycytidine Kinase ◽  
Wide Range ◽  
Substrate Concentrations

Natural ribonucleoside-5’-monophosphates are building blocks for nucleic acids which are used for a number of purposes, including food additives. Their analogues, additionally, are used in pharmaceutical applications. Fludarabine-5´-monophosphate, for example, is effective in treating hematological malignancies. To date, ribonucleoside-5’-monophosphates are mainly produced by chemical synthesis, but the inherent drawbacks of this approach have led to the development of enzymatic synthesis routes. In this study, we evaluated the potential of human deoxycytidine kinase (HsdCK) as suitable biocatalyst for the synthesis of natural and modified ribonucleoside-5’-monophosphates from their corresponding nucleosides. Human dCK was heterologously expressed in E. coli and immobilized onto Nickel-nitrilotriacetic acid (Ni-NTA) superflow. A screening of the substrate spectrum of soluble and immobilized biocatalyst revealed that HsdCK accepts a wide range of natural and modified nucleosides, except for thymidine and uridine derivatives. Upon optimization of the reaction conditions, HsdCK was used for the synthesis of fludarabine-5´-monophosphate using increasing substrate concentrations. While the soluble biocatalyst revealed highest product formation with the lowest substrate concentration of 0.3 mM, the product yield increased with increasing substrate concentrations in the presence of the immobilized HsdCK. Hence, the application of immobilized HsdCK is advantageous upon using high substrate concentration which is relevant in industrial applications.

Calixarene Phosphorylation: Cation Control of Reaction Pathways

1993 ◽  
Vol 46 (11) ◽  
pp. 1673 ◽  
Author(s):  
LT Byrne ◽  
JM Harrowfield ◽  
DCR Hockless ◽  
BJ Peachey ◽  
BW Skelton ◽  
...  
Keyword(s):  
Phase Transfer ◽  
Tetrabutylammonium Bromide ◽  
Crystal Structure Analysis ◽  
Bidentate Ligand ◽  
Tetraethylammonium Bromide ◽  
Cone Conformation ◽  
Formation Reaction ◽  
Partial Cone ◽  
Major Reaction Product ◽  
Major Reaction

The hydroxide ion catalysed phosphorylation of p-t- butylcalix [4] arene by diethyl chlorophosphate under heterogeneous conditions shows a marked sensitivity to the nature of the phase transfer catalyst used. Thus, substitution of tetraethylammonium bromide for the tetrabutylammonium bromide prescribed for the synthesis of the calixarene tetraphosphate leads to a major reaction product becoming a bis (phosphate) in which the calixarene binds as a unidentate ligand to one phosphorus and as a bidentate ligand to the other. An X-ray crystal structure analysis on this new compound shows the calixarene to adopt a somewhat distorted 'cone' conformation with an unusual orientation of one of the t-butyl substituents into the cone cavity. It also reveals that a significant diastereoselectivity must operate in the formation reaction. The only other reaction product detected in this system appears to be the calixarene tetraphosphate in a partial cone conformation.

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