Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.5.1.4 (deaminase)
 5,113 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Formamidase from rat liver proved to be microheterogenous. After preparative isoelectric focusing in density gradient columns, two peaks of formamidase with identical substrate specificity were identified. By analytical focusing in thin layers of polyacrylamide or Sephadex G-75 SF, even five bands could be separated. Their isoelectric points were 4.75, 4.78, 4.82, 4.92 (main band) and 5.11, but their Michaelis constants did not differ significantly (54 to 62 mumol/l). An identical molecular weight of 34700 +/- 3200 for all bands was determined by disc electrophoresis. This value was confirmed by sedimentation analyses (so20,w = 3.00 S) and electrophoresis in the presence of sodium dodecyl-sulfate (Mr 34900 +/- 2300), which only gave a single band. The homogeneity was also confirmed by electrophoresis in the presence of 6M urea. Repeated disc electrophoresis of focusing under native conditions with single, isolated formamidases again resulted in different bands which were identified, not only by Coomassie Blue, but also by their hydrolytic cleavage of naphthyl acetate. Formamidase showed neither proteolytic nor asparagine-amidohydrolase activity and oligosaccharide conjugates were not detectable. Ampholytes, buffer ions, pH and peroxodisulfate did not affect the heterogeneity. "Initial burst" measurements with diethyl(4-nitrophenyl) phosphate yielded an equivalent weight of 36,300. Formylkynurenine reduced this inhibition very effectively. Thus, an extraordinary reactive serine residue appeared to be located in the catalytic site of formamidase. A participation of sulfhydrylgroups in the inactivating reaction of arsenite was excluded although two such groups were detected by 5,5'-dithiobis(2-nitrobenzoic acid). N-Bromosuccinimide reacted primarily with one of the nine tryptophan residues without loss of enzymatic activity, but a 18.6-fold excess of this reagent resulted in a complete loss of activity. The reaction rates of the most effective inhibitors and of the protective action of formylkynurenine were determined. Thus, formamidase must clearly be distinguished from typical serine esterases and proteases.
 ...
PMID:[Formamidase--microheterogeneity, catalytic properties and inhibitors (author's transl)]. 8 81

The Swatek's method was further simplified for the assay of penicillin amidase activity. The absorbance of colour obtained during determination of 6-aminopenicillanic acid was dependent on concentration of 4-dimethylaminobenzaldehyde and on temperature. Antiodies induced in rabbits with one molecular form of penicillin amidase from E. coli PCM 271 (PA-1 or PA-2) did not cross-react with the other amidase form. No differences in substrate specificity on inactivation with SDS and in alkaline medium between the two amidase forms were observed. Concentrated urea inactivated PA-2 irreversibly and PA-1 reversibly. N-Bromosuccinimide inactivated almost completely only PA-1. Two E. coli PCM 271 strain variants were separated by microbial selection. Each of them produced only one amidase form. Also two amidase forms were found in cells of E. coli ATCC 11105, whereas E. coli ATCC 9636 and ATCC 9637 synthesize only PA-1.
 ...
PMID:Two molecular forms of penicillin amidase synthesized by Escherichia coli. 283 78

Penicillin V acylase (PVA), a member of newly evolved Ntn-hydrolase superfamily, is a pharmaceutically important enzyme to produce 6-aminopenicillanic acid. Active site characterization of recently purified monomeric PVA from Rhodotorula aurantiaca (Ra-PVA), the yeast source, showed the involvement of serine and tryptophan in the enzyme activity. Modification of the protein with serine and tryptophan specific reagents such as PMSF and NBS showed partial loss of PVA activity and substrate protection. Ra-PVA was found to be a multi-tryptophan protein exhibiting one tryptophan, in native and, four in its denatured condition. Various solute quenchers and substrate were used to probe the microenvironment of the putative reactive tryptophan through fluorescence quenching. The results obtained indicate that the tryptophan residues of Ra-PVA were largely buried in hydrophobic core of the protein matrix. Quenching of the fluorescence by acrylamide was collisional. Acrylamide was the most effective quencher amongst all the used quenchers, which quenched 71.6% of the total intrinsic fluorescence of the protein, at a very less final concentration of 0.1M. Surface tryptophan residues were found to have predominantly more electropositively charged amino acids around them, however differentially accessible for ionic quenchers. Denaturation led to shift in lambda(max) from 336, in native state, to 357 nm and more exposed to the solvent, consequently increase in fluorescence quenching with all quenchers. This is an attempt towards the conformational studies of Ra-PVA.
 ...
PMID:Rhodotorula aurantiaca penicillin V acylase: active site characterization and fluorometric studies. 1981 16