Water solubility, stability, and bioavailability, can be substantially improved after glycosylation. Glycosylation of bioactive compounds catalyzed by glycoside hydrolases (GHs) and glycosyltransferases (GTs) has become a research hotspot. Thanks to their rich sources and use of cheap glycosyl donors, GHs are advantageous in terms of scaled catalysis compared to GTs. Among GHs, sucrose phosphorylase has attracted extensive attentions in chemical engineering due to its prominent glycosylation activity as well as its acceptor promiscuity. This paper reviews the structure, catalytic characteristics, and directional redesign of sucrose phosphorylase. Meanwhile, glycosylation of diverse chemicals with sucrose phosphorylase and its coupling applications with other biocatalysts are summarized. Future research directions were also discussed based on the current research progress combined with our working experience.
Glucosyltransferases/metabolism* ; Glycoside Hydrolases/metabolism* ; Glycosylation ; Glycosyltransferases/genetics*

The lignan glycosyltransferase UGT236(belonging to the UGT71 B family) from Isatis indigotica can catalyze the production of phloridzin from phloretin in vitro. UGT236 shares high identity with P2'GT from apple. In this study, the recombinant plasmid pET28 a-MBP-UGT236 was transferred into Escherichia coli Rosetta(DE3) cells and induced by isopropyl-β-D-thiogalactoside(IPTG). The purified UGT236 protein was used for enzymatic characterization with phloretin as substrate. The results showed that UGT236 had the optimal reaction temperature of 40 ℃ and the optimal pH 8(Na_2HPO_4-NaH_2PO_4 system). The UGT236 activity was inhibited by Ni~(2+) and Al~(3+), enhanced by Fe~(2+), Co~(2+), and Mn~(2+), and did not affected by Mg~(2+), Ca~(2+), Li~+, Na~+, or K~+. The K_m, K_(cat), and K_(cat)/K_m of phloretin were 61.03 μmol·L~(-1), 0.01 s~(-1), and 157.11 mol~(-1)·s~(-1)·L, and those of UDPG were 183.6 μmol·L~(-1), 0.01 s~(-1), and 51.91 mol~(-1)·s~(-1)·L, respectively. The possible active sites were predicted by homologous modeling and molecular docking. By mutagenisis and catalytic activity detection, three key active sites, Glu391, His15, and Thr141, were identified, while Phe146 was related to product diversity. In summary, we found that the lignan glycosyltransferase UGT236 from I.indigotica could catalyze the reaction of phloretin into phloridzin. Several key amino acid residues were identified by structure prediction, molecular docking, and site-mutagenesis, which provided a basis for studying the specificity and diversity of phloretin glycoside products. This study can provide a reference for artificially producing glycosyltransferase elements with high efficiency and specific catalysis.
Glucosyltransferases/genetics* ; Glycosyltransferases/metabolism* ; Isatis ; Lignans/metabolism* ; Molecular Docking Simulation ; Phloretin/metabolism* ; Phlorhizin/metabolism*

Glycogen storage disease type V (GSD-V) is the most common disorder of muscle glycogenosis with characteristic clinical and laboratory findings. A 32-yr-old woman complained of exercise intolerance and myoglobulinuria since early adolescence. She reported several episodes of second-wind phenomenon. Physical examination did not show any neurological abnormality, including fixed muscle weakness or atrophy. Serum creatine kinase level was 1,161 IU/L at rest. The result of the non-ischemic forearm exercise test was compatible with GSD-V. Mutation analysis identified the compound heterozygous mutations of the PYGM, p.D510fs and p.F710del, which has not yet been reported in Korea. The present case recognizes that detail clinical and laboratory analysis is the first step in the diagnosis of GSD-V.
Adult ; Base Sequence ; Creatine Kinase/blood ; Exons ; Female ; Frameshift Mutation ; Gene Deletion ; Genotype ; Glycogen Phosphorylase, Muscle Form/genetics ; Glycogen Storage Disease Type V/*diagnosis/genetics/pathology ; Humans ; Pedigree ; Sequence Analysis, DNA

The mature cyst of Acanthamoeba is highly resistant to various antibiotics and therapeutic agents. Cyst wall of Acanthamoeba are composed of cellulose, acid-resistant proteins, lipids, and unidentified materials. Because cellulose is one of the primary components of the inner cyst wall, cellulose synthesis is essential to the process of cyst formation in Acanthamoeba. In this study, we hypothesized the key and short-step process in synthesis of cellulose from glycogen in encysting Acanthamoeba castellanii, and confirmed it by comparing the expression pattern of enzymes involving glycogenolysis and cellulose synthesis. The genes of 3 enzymes, glycogen phosphorylase, UDP-glucose pyrophosphorylase, and cellulose synthase, which are involved in the cellulose synthesis, were expressed high at the 1st and 2nd day of encystation. However, the phosphoglucomutase that facilitates the interconversion of glucose 1-phosphate and glucose 6-phosphate expressed low during encystation. This report identified the short-cut pathway of cellulose synthesis required for construction of the cyst wall during the encystation process in Acanthamoeba. This study provides important information to understand cyst wall formation in encysting Acanthamoeba.
Acanthamoeba castellanii/*enzymology/genetics/growth & development ; Amebiasis/*parasitology ; Cell Wall/*metabolism ; Cellulose/*biosynthesis/genetics ; Glucosyltransferases/genetics/metabolism ; Glycogen Phosphorylase/genetics/metabolism ; Protozoan Proteins/genetics/*metabolism ; UTP-Glucose-1-Phosphate Uridylyltransferase/genetics/metabolism

In this study, PCR technique was used with 2 primer pairs for amplifying DNA fragments at 5’ and 3’ of the gene encoding diphteria toxine of the strains Corynebacterium diphtheriae. After the cloning and sequencing, 2 separated fragments were joined to form a complete gene. The sequence of gene was translated into protein and the results were submitted into Gene Bank Database
Diphtheria Toxin ; Genes ; Corynebacterium diphtheriae

Aims: Cholera epidemics have been occurred in Malaysia since 1991 till 2003 which can be proved from the records by the Infectious Diseases Division of the Ministry of Health. Moreover, there were also course of cholera epidemics from the year 1994 to 2003 which had been happened in Sarawak. Cholera outbreaks in Malaysia mostly caused by the El Tor O1 Vibrio cholerae serogroup. The aims of this study were to detect the presence of V. cholerae in clinical and environmental samples (n=28) from Limbang, Sarawak by collaboration with Sarawak Government Hospital and to detect the toxin genes from the isolates. Methodology and results: All the isolates were sub-cultured in alkaline peptone water (APW). The boiled-cell method was used for DNA extraction. The total DNA extracted was amplified by polymerase chain reaction (PCR). Two types of PCR were used in this study which are 16S rRNA PCR and multiplex PCR. The results obtained from the study found out that 16 out of 28 (57.14%) samples were confirmed to be V. cholerae species. Four primers specific for V. cholerae were used in multiplex PCR (O1 type, O139 type, ctxA and ctxAB) to confirm the species type and the toxin genes. All samples shown positive for V. cholerae O1 serotype and 100% positive to all genes for the identification of ctxA and ctxAB genes. Conclusion, significance and impact of study From this study, it showed that multiplex PCR can be used for research purposes in molecular genetics field involving cholera outbreak.
Vibrio cholerae--genetics ; Cholera Toxin

Here we have investigated how lactosylceramide (LacCer) modulates gene expression of adhesion molecules in TNF-alpha and IFNgamma (CM)-stimulated astrocytes. We have observed that stimulation of astrocytes with CM increased the gene expression of ICAM-1 and VCAM-1. D-Threo-1-phenyl- 2-decanoylamino-3-morpholino-1-propanol (PDMP) and N-butyldeoxynojirimycin (NBDNJ), inhibitors of glucosylceramide synthase (GLS) and LacCer synthase (galactosyltransferase, GalT-2), inhibited the gene expression of ICAM-1 and VCAM-1 and activation of their gene promoter induced by CM, which were reversed by exogenously supplied LacCer. Silencing of GalT-2 gene using its antisense oligonucleotides also attenuated CM-induced ICAM-1 and VCAM-1 expression, which were reversed by LacCer. PDMP treatment and silencing of GalT-2 gene significantly reduced CM-induced luciferase activities in NF-KB, AP-1, GAS, and STAT-3 luciferase vectors-transfected cells. In addition, LacCer reversed the inhibition of NF-KB and STAT-1 luciferase activities by PDMP. Taken together, our results suggest that LacCer may play a crucial role in the expression of ICAM-1 and VCAM-1 via modulating transcription factors, such as NF-KB, AP-1, STAT-1, and STAT-3 in CM-stimulated astrocytes.
1-Deoxynojirimycin ; Antigens, CD ; Astrocytes ; Galactosyltransferases ; Gene Expression ; Glucosyltransferases ; Intercellular Adhesion Molecule-1 ; Lactosylceramides ; Luciferases ; Morpholines ; NF-kappa B ; Oligonucleotides, Antisense ; Transcription Factor AP-1 ; Transcription Factors ; Tumor Necrosis Factor-alpha ; Vascular Cell Adhesion Molecule-1

No abstract available.
Calcium* ; Cholera Toxin* ; Cholera* ; Cytoplasm* ; Humans* ; T-Lymphocytes*

We used cholera toxin B subunit (CTB) as a neural tracer to localize motor neuronal cell bodies innervating the digastric muscle. After CTB injection into the left anterior belly, CTB-labelled motor neuronal cell bodies were found in caudal half of the left and right trigeminal nucleus, the left and right facial nucleus, the accessory facial nucleus and the accessory trigeminal nucleus in pons. The total number of CTB-labelled motor neuronal cell bodies were 1,179+/-119.5 in the left pons and 246+/-61.8 in the right pons after CTB injections into the left anterior belly of digastric muscle. After CTB injection into left posterior belly, CTB-labelled motor neuronal cell bodies were found only in the left ventral part of accessory facial nucleus in caudal pons and the total number of CTB-labelled motor neuronal cell bodies were 270+/-29.3.
Animals ; Cholera Toxin* ; Cholera* ; Motor Neurons* ; Pons ; Rats* ; Trigeminal Nuclei

No abstract available.
Animals ; Glycogen Synthase ; Glycogen ; Insulin ; Rats