scholarly journals Purification and some properties of an IMP-specific 5′-nucleotidase from yeast

1994 ◽  
Vol 298 (3) ◽  
pp. 593-598 ◽  
Author(s):  
R Itoh
Keyword(s):  
Molecular Mass ◽  
Substrate Concentration ◽  
Soluble Protein ◽  
The Other ◽  
Metal Salts ◽  
Bivalent Metal ◽  
Pyrimidine Nucleotides ◽  
Ph Optimum ◽  
A Subunit ◽  
Purine And Pyrimidine Nucleotides

An IMP-hydrolysing enzyme was purified to homogeneity from yeast extract. It was a soluble protein with an apparent molecular mass of 220 kDa, with a subunit molecular mass of 55 kDa. It was highly specific for IMP, and there was virtually no detectable activity with the other purine and pyrimidine nucleotides tested, including AMP and dIMP. The enzyme had a pH optimum of 6.0-6.5. Its activity was absolutely dependent on bivalent metal salts: Mg2+ was most potent, followed by Co2+ and Mn2+. The velocity/substrate-concentration plot of the enzyme was slightly sigmoidal (h = 1.7) and the s0.5 was 0.4 mM. ATP stimulated the enzyme by decreasing both h and s0.5. Diadenosine tetraphosphate stimulated the enzyme as effectively as ATP. Although the properties of the enzyme are similar to those of the IMP/GMP 5′-nucleotidase identified in various animals [Itoh (1993) Comp. Biochem. Physiol. 105B, 13-19], the substrate specificity of the former was much more strict than the latter.

scholarly journals The deoxyribonucleoside phosphotransferase of Trichomonas vaginalis. A potential target for anti-trichomonial chemotherapy.

1984 ◽  
Vol 160 (4) ◽  
pp. 987-1000 ◽  
Author(s):  
C C Wang ◽  
H W Cheng
Keyword(s):  
Trichomonas Vaginalis ◽  
De Novo ◽  
Protozoan Parasite ◽  
Pyrimidine Nucleotides ◽  
Ph Optimum ◽  
Potential Target ◽  
Competitive Inhibitors ◽  
Purine And Pyrimidine Nucleotides ◽  
Triton X ◽  
Enzyme I

Trichomonas vaginalis, a human protozoan parasite known to lack the capability of synthesizing purine and pyrimidine nucleotides de novo, was found also incapable of converting its ribonucleotides to deoxyribonucleotides. The only apparent means of providing deoxyribonucleotides for DNA synthesis relies on salvaging exogenous deoxyribonucleosides by a deoxyribonucleoside phosphotransferase activity in the T. vaginalis 10(5) g pelletable fraction. The activity, constituted by at least two isozymes I and II, can be solubilized by Triton X-100, has a pH optimum of 5.0-6.0, and recognizes only thymidine, deoxyadenosine, deoxyguanosine, and deoxycytidine as the phosphate acceptor. TMP, dAMP, dGMP, dCMP, dUMP, FdUMP, and p-nitrophenylphosphate can serve as phosphate donors. Enzyme I has been purified 10-fold by DEAE-Sepharose chromatography and Sephacryl 200 filtration, and is totally freed of the acid phosphatase of T. vaginalis. It has an estimated molecular weight of 200,000 and Km values of 2-3 mM for the four deoxyribonucleosides, which act on each other as competitive inhibitors. It also possesses phosphatase activity capable of hydrolyzing p-nitrophenylphosphate with a Michaelis constant of 0.74 mM. The rates of hydrolysis are enhanced by thymidine, which suggests that the latter may be the preferred phosphate acceptor, and Enzyme I may be, thus, more a transferase than a phosphatase. This enzyme could be a potential target for antitrichomonial chemotherapy.

Purification of tributyrin esterase fromLactococcus lactissubsp.cremorisE8

1996 ◽  
Vol 63 (1) ◽  
pp. 131-140 ◽  
Author(s):  
Ross Holland ◽  
Tim Coolbear
Keyword(s):  
Molecular Mass ◽  
Lactococcus Lactis ◽  
Half Life ◽  
Esterase Activity ◽  
Terminal Sequence ◽  
Ph Optimum ◽  
Single Protein ◽  
A Subunit

SummaryA tributyrin esterase was purified fromLactococcus lactissubsp.cremorisE8 using FPLC chromatography. This was the major esterase activity observed in strain E8 and was associated with a single protein with a subunit molecular mass of 29 kDa and a holoenzyme of molecular mass 109 kDa. The enzyme was active against tributyrin andp-nitrophenyl butyrate. The N-terminal sequence of the enzyme was determined. The enzyme had a pH optimum in the neutral range, was stable on freezing at −20 °C, and had a half life of 1 h at 50 °C.

scholarly journals Control of Drosophila deoxyuridine triphosphatase. Existence of a developmentally expressed protein inhibitor

1989 ◽  
Vol 259 (2) ◽  
pp. 593-596 ◽  
Author(s):  
M D Nation ◽  
S N Guzder ◽  
L E Giroir ◽  
W A Deutsch
Keyword(s):  
Drosophila Melanogaster ◽  
Molecular Mass ◽  
Sedimentation Coefficient ◽  
The Other ◽  
Proteinase K ◽  
Protein Inhibitor ◽  
Stages Of Development ◽  
Early Stages ◽  
A Subunit ◽  
Rnase Treatment

An activity that inhibits deoxyuridine triphosphatase (dUTPase) has been partially purified from Drosophila melanogaster. The inhibitor has a sedimentation coefficient of 4.1 S and a subunit molecular mass of 61 kDa. Its expression is limited to early stages of development, similar to the pattern previously found for dUTPase. The inhibitor is unusually stable to heating and is insensitive to DNAse and RNAse treatment. On the other hand, inhibition is sensitive to digestion with proteinase K, indicating that a protein is required for activity. These results suggest that at least one form of regulation is exerted on Drosophila dUTPase that could allow a greater opportunity for the incorporation of uracil into DNA.

scholarly journals β-Fructofuranosidases from roots of dandelion (Taraxacum officinale Weber)

1972 ◽  
Vol 126 (3) ◽  
pp. 569-573 ◽  
Author(s):  
P. P. Rutherford ◽  
A. C. Deacon
Keyword(s):  
General Formula ◽  
Soluble Protein ◽  
Hydrolytic Enzymes ◽  
Deae Cellulose ◽  
Physiological Role ◽  
Taraxacum Officinale ◽  
The Other ◽  
Ph Optimum ◽  
Cellulose Column

1. Three β-fructofuranosidases were separated by chromatography on a DEAE-cellulose column from the soluble protein extracted from dandelion (Taraxacum officinale Weber) roots. 2. One enzyme, which acted on sucrose, was characterized as an invertase, with a Km of 2.00×10-2M and pH optimum of 7.5. 3. The other two enzymes are hydrolases (A and B), which act on the inulin series of oligosaccharides [general formula glucose-fructose-(fructose)n]. They both have a pH optimum of 4.0 and Km of 1.54×10-2M but differ in their chromatographic behaviour on DEAE-cellulose. Neither of the hydrolases is inhibited by sucrose. 4. The physiological role of these three hydrolytic enzymes is discussed.

scholarly journals UDP-glucose:solasodine glucosyltransferase from eggplant (Solanum melongena L.) leaves: partial purification and characterization.

1997 ◽  
Vol 44 (1) ◽  
pp. 43-53 ◽  
Author(s):  
C Paczkowski ◽  
M Kalinowska ◽  
Z A Wojciechowski
Keyword(s):  
Molecular Mass ◽  
Solanum Melongena ◽  
Divalent Metal ◽  
Partial Purification ◽  
Divalent Metal Ions ◽  
Purification And Characterization ◽  
Ph Optimum ◽  
Cytosol Fraction ◽  
Ammonium Sulphate Precipitation ◽  
Unripe Fruits

Uridine 5'-diphosphoglucose-dependent glucosyltransferase which catalyzes the glycosylation of solasodine i.e. UDP-glucose:solasodine glucosyltransferase, is present in leaves, roots, unripe fruits and unripe seeds of eggplant (Solanum melongena L.). The glucosylation product is chromatographically identical with authentic solasodine 3 beta-D-monoglucoside, a putative intermediate in the biosynthesis of solasodine-based glycoalkaloids characteristic of the eggplant. The enzyme was purified about 50-fold from crude cytosol fraction of eggplant leaves by ammonium sulphate precipitation and column chromatography on Q-Sepharose and Sephadex G-100. The native enzyme has a molecular mass of approx. 55 kDa and pH optimum of 8.5. Divalent metal ions are not required for its activity but the presence of free-SH groups is essential. Besides solasodine (Km = 0.04 microM), the enzyme effectively glucosylates tomatidine, another steroidal alkaloid of the spirosolane type, but it is virtually inactive towards the solanidane-type steroidal alkaloids such as solanidine or demissidine. The enzyme is specific for UDP-glucose (Km = 2.1 microM) since unlabelled ADP-, GDP-, CDP- or TDP-glucose could not effectively compete with UDP-[14C]glucose used as the sugar donor for solasodine glucosylation. Moreover, no synthesis of labelled solasodine galactoside was observed when UDP-[14C]glucose was replaced with UDP-[14C]galactose.

Physicochemical properties of a novel α-L-arabinofuranosidase fromRhizomucor pusillusHHT-1

2001 ◽  
Vol 47 (8) ◽  
pp. 767-772 ◽  
Author(s):  
A KM Shofiqur Rahman ◽  
Shinya Kawamura ◽  
Masahiro Hatsu ◽  
M M Hoq ◽  
Kazuhiro Takamizawa
Keyword(s):  
Amino Acid ◽  
Amino Acid Sequence ◽  
Molecular Mass ◽  
Gel Filtration ◽  
Hydrolytic Activity ◽  
Exchange Chromatography ◽  
Ph Optimum ◽  
Rhizomucor Pusillus ◽  
Terminal Amino ◽  
Metal Cofactors

The zygomycete fungus Rhizomucor pusillus HHT-1, cultured on L(+)arabinose as a sole carbon source, produced extracellular α-L-arabinofuranosidase. The enzyme was purified by (NH4)2SO4fractionation, gel filtration, and ion exchange chromatography. The molecular mass of this monomeric enzyme was 88 kDa. The native enzyme had a pI of 4.2 and displayed a pH optimum and stability of 4.0 and 7.0–10.0, respectively. The temperature optimum was 65°C, and it was stable up to 70°C. The Kmand Vmaxfor p-nitrophenyl α-L-arabinofuranoside were 0.59 mM and 387 µmol·min–1·mg–1protein, respectively. Activity was not stimulated by metal cofactors. The N-terminal amino acid sequence did not show any similarity to other arabinofuranosidases. Higher hydrolytic activity was recorded with p-nitrophenyl α-L-arabinofuranoside, arabinotriose, and sugar beet arabinan; lower hydrolytic activity was recorded with oat–spelt xylan and arabinogalactan, indicating specificity for the low molecular mass L(+)-arabinose containing oligosaccharides with furanoside configuration.Key words: α-L-arabinofuranosidase, enzyme purification, amino acid sequence, Rhizomucor pusillus.

Sign in / Sign up

Export Citation Format

Share Document