• Journal of Microbiology and Biotechnology
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• v.26 no.11
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• pp.1845-1854
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• 2016

The transglycosylation activity of amylosucrase (ASase) has received significant attention owing to its use of an inexpensive donor, sucrose, and broad acceptor specificity, including glycone and aglycone compounds. The transglycosylation reaction of recombinant ASase from Deinococcus radiopugnans (DRpAS) was investigated using various phenolic compounds, and quercetin-3-O-rutinoside (rutin) was found to be the most suitable acceptor molecule used by DRpAS. Two amino acid residues in DRpAS variants (DRpAS Q299K and DRpAS Q299R), assumed to be involved in acceptor binding, were constructed by site-directed mutagenesis. Intriguingly, DRpAS Q299K and DRpAS Q299R produced 10-fold and 4-fold higher levels of rutin transglycosylation product than did the wild-type (WT) DRpAS, respectively. According to in silico molecular docking analysis, the lysine residue at position 299 in the mutants enables rutin to more easily position inside the active pocket of the mutant enzyme than in that of the WT, due to conformational changes in loop 4.

• Korean Journal of Microbiology
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• v.35 no.2
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• pp.146-152
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• 1999

Acceplor reaction of glucosyltr~ansferase of Streptococcus ,SO~~-~IZLIS with f ~ ~ t o ~ y l o l i g o ~ a ~ ~ h a r i d e ~ was studied for the biosynthesis of novel olgisaccharides. First, bacilracin resistant mutants were selected by mutagenesis of Streptococcus sobrimis ATCC27351. Among these mutants 4 strains were selected by resistance to bacitracin and increase of glucosyltransferase. Acceptor reaction of maltose was analyzed by TLC and image analysis. There were differences in the specificity of the acceptor reaclion by Ule glucosylumsferase between mother strain (Streptococcus sobrinus ATCC2735) and bacitracin resistant mutants (Streptococcus sobrinus BR24C, Strepfococcus sobrinus CH-5). Molher strain did ilot show an acceptor reaction with fructosyloligosaccharides such as 1-keqtose and nystose. Acceptor reaction products of turailose and 1-kestose with glucosyltransferase (GW-S) of Streptococcus sobrini~s BR24C were TEX>\6^{3}$-$\alpha$-D-glucopyranosyl \3^{2}$-O-$\alpha$-D-fructose (glucose-fructose-glucose) and \6^{4}$-$\alpha$-D-glucopyranosyl \1^{3}$-$\beta$-D-~-h~ctofuranos~~ sucrose (glucose-glucosefructose- fructose). respectively These are novel oligosaccharides which can be produced only by enzymatic reaction.

• Journal of Microbiology and Biotechnology
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• v.15 no.1
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• pp.99-104
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• 2005

Microbacterium laevaniformans levansucrase synthesized various hetero-oligosaccharides by transferring fructosyl residue from sucrose to various saccharides as acceptors. The acceptor specificity test showed that reducing saccharides were more favorable acceptors than nonreducing saccharides. The transfructosylated product, fructosyl galactose, was produced in the presence of D-galactose as an acceptor. The chemical structure of the resulting fructosyl galactose was analyzed by yeast invertase and NMR, and identified as O-$\alpha$-D-galactosyl-(1${\to}$2)-$\beta$-D-fructofuranoside. These results indicate that the main transfructosylation activity of the enzyme is to make nonreducing transferred products via a transfer of fructosyl residue to acceptor molecules having reducing group. When nonreducing sugars, such as methyl $\alpha$-D-glucoside and methyl $\alpha$-D-galactoside, were used as an acceptor, the transfer product was also formed in spite of the reducing group blocked with methyl group. The fact that no transfer product was formed with sugar alcohols as acceptors was suggested to be due to marked conformational difference of acceptors.

• Bulletin of the Korean Chemical Society
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• v.18 no.1
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• pp.18-23
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• 1997

Data from structure/activity studies in vir gene induction system have led to evaluate the working hypothesis of interaction between phenolic inducers and phenol binding proteins. The primary specificity in the association of a phenolic inducer with its receptor in our system is hypothesized to be the hydrogen bonding interactions through the ortho methoxy substituents as well as the proton transfer between the inducer and the binding protein. In this paper the proposed working model for phenol-mediating signal transduction was evaluated in several ways. The importance of the general acid-base catalysis was first addressed by the presence of an acidic residue and a basic residue in the phenol binding protein. Series of compounds were tested for vir gene expression activity to confirm the generation of a strong nucleophile by an acidic residue and an involvement of a basic residue as a proton acceptor. An attempt was made to correlate the pKa values of the phenolic compounds with vir gene induction activities as inducers to further support the proposed proton transfer mechanism. Finally, it was also observed that the regioselectively attached methoxy group on phenol compounds is required as the proper hydrogen bond acceptor.

• Proceedings of the Korean Biophysical Society Conference
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• 1997.07a
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• pp.37-37
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• 1997

The catalytic efficiency of pyranose oxidase (EC 1.1.3.10.) determined for various sugars showed that D-glucose is the preferred substrate and the enzyme oxidized the various aldonolactones. The specificity constants of pyranose oxidase determined for deoxy- and deoxyfluoro-D-glucoses showed that a hydroxy group at C-4 of D-glucose acts as a hydrogen-bone acceptor, at C-6 as a hydrogen-bond donor, and at C-1 as a hydrogen-bond donor.(omitted)

• Applied Biological Chemistry
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• v.21 no.2
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• pp.112-118
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• 1978

Xanthine oxidase from bovine thyroid glands was purified to apparent homogeneity when judged by analytical disc gel electrophoresis. The purification procedures include pancreatin digestion, butanol extraction, ammonium sulfate precipitation, calcium phosphate gel adsorption, ultrafiltration, calcium phosphate gel-cellulose column chromatography, gel filtration, preparative Sephadex G-25 column electrophoresis, and preparative polyacrylamide gel electrophoresis. The enzyme was enriched 1,000-fold. However, its specific activity was markedly low as compared with highly purified milk enzyme. Thyroidal xanthine oxidase exhibited a low specificity for substrates and electron acceptors. The kinetic properties of thyroid xanthine oxidase were found to be similar to those of the milk enzyme on the basis of Michaelis constants for common substrates.

• Journal of Microbiology and Biotechnology
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• v.29 no.2
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• pp.268-273
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• 2019

The specificity of a Bacillus licheniformis uridine diphosphate (UDP) glycosyltransferase, YjiC, was increased towards thymidine diphosphate (TDP)-sugar by site-directed mutagenesis. The Arg-282 of YjiC was identified and investigated by substituting with Trp. Conversion rate and kinetic parameters were compared between YjiC and its variants with several acceptor substrates such as 7-hydroxyflavone (7-HF), 4',7-dihydroxyisoflavone, 7,8-dihydroxyflavone and curcumin. Molecular docking of TDP-glucose and 7-HF with YjiC model showed pi-alkyl interaction with Arg-282 and His-14, and pi-pi interaction with $His^{14}$ and thymine ring. YjiC (H14A) variant lost its glucosylation activity with TDP-glucose validating significance of His-14 in binding of TDP-sugars.

• Journal of Microbiology and Biotechnology
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• v.30 no.9
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• pp.1436-1442
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• 2020

Amylosucrase (ASase, E.C. 2.4.1.4) is capable of efficient glucose transfer from sucrose, acting as the sole donor molecule, to various functional acceptor compounds, such as polyphenols and flavonoids. An ASase variant from Deinococcus geothermalis, in which the 226^th alanine is replaced with asparagine (DgAS-A226N), shows increased polymerization activity due to changes in the flexibility of the loop near the active site. In this study, we further investigated how the mutation modulates the enzymatic activity of DgAS using molecular dynamics and docking simulations to evaluate interactions between the enzyme and phenolic compounds. The computational analysis revealed that the A226N mutation could induce and stabilize structural changes near the substrate-binding site to increase glucose transfer efficiency to phenolic compounds. Kinetic parameters of DgAS-A226N and WT DgAS were determined with sucrose and 4-methylumbelliferone (MU) as donor and acceptor molecules, respectively. The k_cat/K_m value of DgAS-A226N with MU (6.352 mM^-1min^-1) was significantly higher than that of DgAS (5.296 mM^-1min^-1). The enzymatic activity was tested with a small phenolic compound, hydroquinone, and there was a 1.4-fold increase in α-arbutin production. From the results of the study, it was concluded that DgAS-A226N has improved acceptor specificity toward small phenolic compounds by way of stabilizing the active conformation of these compounds.

• Journal of Microbiology and Biotechnology
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• v.17 no.3
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• pp.454-460
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• 2007

Enzymatic properties and substrate specificity of recombinant $\beta-glycosidases$ from a hyperthermophilic archaeon, Sulfolobus shibatae (rSSG), were analyzed. rSSG showed its optimum temperature and pH at $95^{\circ}C$ and pH 5.0, respectively. Thermal inactivation of rSSG showed that its half-life of enzymatic activity at $75^{\circ}C$ was 15 h whereas it drastically decreased to 3.9 min at $95^{\circ}C$. The addition of 10 mM of $MnCl_2$ enhanced the hydrolysis activity of rSSG up to 23% whereas most metal ions did not show any considerable effect. Dithiothreitol (DTT) and 2-mercaptoethanol exhibited significant influence on the increase of the hydrolysis activity of rSSG rSSG apparently preferred laminaribiose $(\beta1\rightarrow3Glc)$, followed by sophorose $(\beta1\rightarrow2Glc)$, gentiobiose $(\beta1\rightarrow6Glc)$, and cellobiose $(\beta1\rightarrow4Glc)$. Various. intermolecular transfer products were formed by rSSG in the lactose reaction, indicating that rSSG prefers lactose as a good acceptor as well as a donor. The strong intermolecular transglycosylation activity of rSSG can be applied in making functional oligosaccharides.

• BMB Reports
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• v.35 no.5
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• pp.513-517
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• 2002

We describe a new method for identifying the sequences that signal the start of translation, and the boundaries between exons and introns (donor and acceptor sites) in human mRNA. According to the mandatory keyword, ORGANISM, and feature key, CDS, a large set of standard data for each signal site was extracted from the ASCII flat file, gbpri.seq, in the GenBank release 108.0. This was used to generate the scoring matrices, which summarize the sequence information for each signal site. The scoring matrices take into account the independent nucleotide frequencies between adjacent bases in each position within the signal site regions, and the relative weight on each nucleotide in proportion to their probabilities in the known signal sites. Using a scoring scheme that is based on the nucleotide scoring matrices, the method has great sensitivity and specificity when used to locate signals in uncharacterized human genomic DNA. These matrices are especially effective at distinguishing true and false sites.