Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.2.1.23 (beta-galactosidase)
 14,648 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We report an enzyme immunoassay procedure for methotrexate measurement that takes less than 3 h to perform. beta-D-Galactosidase (EC 3.2.1.23) from Escherichia coli was conjugated to methotrexate by means of the mixed anhydride reaction. Bound and free labeled drug were separated by a preincubated cubic complex of first and second antibody. The enzyme activity of the bound fraction was measured with o-nitrophenyl-beta-D-galactopyranoside as substrate. The standard curve covered the range 1 to 10 micrograms of methotrexate per liter. One microgram of methotrexate per liter inhibited binding of the tracer by 17%. The assay is specific for methotrexate in the presence of folinic acid (citrovorum factor), folic acid, tetrahydrofolic acid, and other methotrexate metabolities. Intra- and inter-assay CVs were less than 5 and 10%, respectively. Results obtained with this enzyme immunoassay method agreed well with those obtained with an established radioimmunoassay method.
 ...
PMID:Improved double-antibody enzyme immunoassay for methotrexate. 11 Apr 99

A hybrid glycinamide ribonucleotide transformylase was assembled from two protein domains that were treated as discrete modules. One module contained the ribonucleotide binding domain from the purN glycinamide ribonucleotide transformylase; the second module contained the catalytic machinery and the formyl tetrahydrofolate binding domain from the enzyme encoded by purU, formyl tetrahydrofolate hydrolase. The resultant enzyme showed 0.1% catalytic activity of the wild-type glycinamide ribonucleotide transformylase enzyme but had a formyl transfer efficiency of 10%. A combinatorial mutagenesis approach was used to improve the solubility and formyl transfer properties of the hybrid enzyme. The mutagenized hybrid glycinamide ribonucleotide transformylase was initially expressed as a fusion to the alpha-peptide of beta-galactosidase. Clones were selected for improvement in solubility by determining which clones were capable of alpha-complementation using a blue/white screen. One clone was further characterized and found to have an improved efficiency of transfer of the ribonucleotide increasing this property to >95%.
 ...
PMID:Improvement in the efficiency of formyl transfer of a GAR transformylase hybrid enzyme. 1083 5

Beta-(1-->4)-thiodisaccharides formed by a pentopyranose unit as reducing or non reducing end have been synthesized using a sugar enone derived from a hexose or pentose as Michael acceptor of a 1-thiopentopyranose or 1-thiohexopyranose derivatives. Thus, 2-propyl per-O-acetyl-3-deoxy-4-S-(beta-D-Xylp)-4-thiohexopyranosid-2-ulose (3) and benzyl per-O-acetyl-3-deoxy-4-S-(beta-D-Galp)-4-thiopentopyranosid-2-ulose (11) were obtained in almost quantitative yields. The carbonyl function of these uloses was reduced with NaBH(4) or K-Selectride, and the stereochemical course of the reduction was highly dependent on the reaction temperature, reducing agent and solvent. Unexpectedly, reduction of 3 with NaBH(4)-THF at 0 degrees C gave a 3-deoxy-4-S-(beta-D-Xylp)-4-thio-alpha-D-ribo-hexopyranoside derivative (6) as major product (74% yield), with isomerization of the sulfur-substituted C-4 stereocenter of the pyranone. Reduction of 11 gave always as major product the benzyl 3-deoxy-4-S-(Galp)-4-thio-beta-D-threo-pentopyranoside derivative 14, which was the only product isolated (80% yield) in the reduction with K-Selectride in THF at -78 degrees C. Deprotection of 14 and its epimer at C-2 (13) afforded, respectively the free thiodisaccharides 19 and 18. They displayed strong inhibitory activity against the beta-galactosidase from Escherichia coli. Thus, compound 18 proved to be a non-competitive inhibitor of the enzyme (K(i)=0.80 mM), whereas 19 was a mixed-type inhibitor (K(i)=32 microM).
 ...
PMID:Synthesis of pentopyranosyl-containing thiodisaccharides. Inhibitory activity against beta-glycosidases. 1967 8