摘要:

The molecular weight of fumarylacetoacetate fumarylhydrolase (EC 3.7.1.2) is 86 000 ± 10 000, as determined by gel filtration. The enzyme appears to be a dimer with a monomer molecular weight of 38 000 – 43 000, as determined by gel electrophoresis, gel filtration in guanidine–hydrochloride, and ultracentrifugation. The subunits appear to be identical, as only one band is seen in gel electrophoresis, only one protein peak is detected in gel filtration in guanidine–hydrochloride, and only one amino-terminal amino acid (proline) is detected. Three free sulfhydryl groups per denatured monomer are detected by reaction with 5,5′-dithiobis(2-nitrobenzoic acid), while for the active enzyme only two sulfhydryl groups react with this reagent. The extinction coefficients at 260 and 280 nm, the amino acid composition, and the isoelectric point (6.7) of the enzyme are also reported.The enzyme catalyzes the hydrolysis of six 2,4-diketo acids and three 3,5-diketo acids tested. TheKmof the substrates is similar butVvaries by a factor of 120. The pH optimum is 7.3. The enzyme did not catalyze the hydrolysis of a number of esters tested.

ISSN:0008-4018    

DOI:10.1139/o77-001

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

The content of histones in transcriptionally active (euchromatin-like (E)) and transcriptionally less active (heterochromatin-like (H)) fractions from the rat ventral prostate was determined. The absolute amount of total histone was less in the E fraction, which contained relatively more stained histone F2band F3and less histone F1than either the H fraction or unfractionated chromatin.The phosphotungstic-acid-stained E fraction was composed predominantly of shorter (50–500 nm) and thinner (5–15 nm) structures. In some preparations, pieces estimated to be 500–1000 + nm in length were present. Spherical structures, from 10 to 20 nm in diameter, some of which were present as 'dimers,' trimers,' or higher-ordered ensembles were seen in the shadow-cast E fraction. The stained or shadow-cast H fraction contained longer (200–2000 + nm) and thickerfragments with a more complex substructure. When the shadow-cast H fraction was sufficiently dispersed, it had a nodular or 'beaded' appearance.

ISSN:0008-4018    

DOI:10.1139/o77-002

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

Cellobiase has been isolated from the crude cellulase mixture of enzymes ofTrichoderma virideusing column chromatographic and ion-exchange methods. The steady-state kinetics of the hydrolysis of cellobiose have been investigated as a function of cellobiose and glucose concentrations, pH of the solution, temperature, and dielectric constant, using isopropanol–buffer mixtures. The results show that (i) there is a marked activation of the reaction by initial glucose concentrations of 4 × 10−3 Mto 9 × 10−2 Mand strong inhibition of the reaction at higher initial concentrations, (ii) the log rate – pH curve has a maximum at pH 5.2 and enzyme pKvalues of 3.5 and 6.8, (iii) the energy of activation at pH 5.1 is 10.2 kcal mol−1over the temperature range 5–56 °C, and (iv) the rate decreases from 0 to 20% (v/v) isopropanol.The hydrolysis by cellobiase (EC 3.2.1.21) ofp-nitrophenyl-β-D-glucoside was examined by pre-steady-state methods in which, and by steady-state methods as a function of pH and temperature. The results show (i) a value fork2of 21 s−1at pH 7.0 (wherek2is the rate constant for the second step in the assumed two-intermediate mechanism) (ii) a log rate–pH curve, significantly different from that for hydrolysis of cellobiose, in which the rate increases with decreasing pH below pH 4.5, is constant in the region pH 4.5–6, and decreases above pH 6 (exhibiting an enzyme pKvalue of 7.3), and (iii) an activation energy of 12.5 kcal mol−1at pH 5.7 over the temperature range 10–60 °C.

ISSN:0008-4018    

DOI:10.1139/o77-003

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

Gigartinine, 5-(3-amidinoureido)-2-aminovaleric acid, andL-citrullinyl-L-arginine were islated from aqueous extracts ofChondrus crispus(Rhodophyceae). Their identifications were confirmed by chemical procedures, and1H and13C nuclear magnetic resonance and infrared spectroscopic methods. Citrullinylarginine, gigartinine, taurine, citrulline, and glutamic acid were the predominant free amino compounds. Citrullinylarginine showed the most pronounced change in concentration. This occurred during the winter months when it reached a maximum (in March) of 58 μmol/g fresh weight, a value 10 times greater than that of any of the free amino acids and equal to 50% of the total organic nitrogen in the plant. All of these compounds were depleted to a minimum of about 1 μmol/g fresh weight in October. Both gigartinine and citrullinylarginine were detected inAhnfeltia plicata,Gracilariasp.Petrocelis middendorfii,Polyides rotundus,Polysiphonia lanosa, andRhodomela confervoides.

ISSN:0008-4018    

DOI:10.1139/o77-004

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

When rat diaphragm, intact or homogenized, was incubated with [14C]glucose, the initial specific activity of the fructose 6-phosphate was higher than that of glucose 6-phosphate (glc-6-P). This relationship was also found when the cytosol was incubated with the labeled glucose. The early time course of production of glc-6-P from glucose and its removal provide evidence that newly synthesized glc-6-P does not equilibrate rapidly with the total pool of this metabolite and is preferentially removed by isomerization. It is concluded that glc-6-P is functionally compartmented within the cytoplasm.

ISSN:0008-4018    

DOI:10.1139/o77-005

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

Sheep β-lipotropin (β-LPH) (sequence 1–91) was selectively cleaved with trypsin after blocking the ϵ-amino groups of lysine with citraconic anhydride. The resulting peptides were purified by a combination of cation-exchange chromatography and high-voltage electrophoresis. The purified fragments were then tested for their morphine-like activity in the mouse vas deferens bioassay. The active peptides were 61–91 and 61–80. All other regions of the molecule investigated were not active. Moreover, the peptides 61–91 and 61–80 were about as active as the synthetic methionine-enkephalin, and in turn these were about 100 times more active than β-LPH itself. The inhibition of electrically stimulated mouse vas deferens by these peptides is reversed by naloxone, and suggests a competitive character of interaction. It is thus concluded that the active core for the morphine-like activity in the mouse vas deferens bioassay is the fragment 61–65 of β-LPH.

ISSN:0008-4018    

DOI:10.1139/o77-006

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

Homogenates of liver and kidney tissue from young steers had β-D-glucosidase (EC 3.2.1.21) activity toward 17α-estradiol-3-glucoside, 17β-estradiol-3-glucoside, 17α-estradiol-17-glucoside, and deoxycorticosterone-21-glucoside. The activity towards the phenolic 3-glucosides was largely present in the 100 000 × gsupernatant, while that towards 17α-estradiol-17-glucoside was concentrated in the microsomes. The use of beef liver preparations for the hydrolysis of steroid 'glucuronide' fractions could result in hydrolysis of other steroid glycosides which might be present.

ISSN:0008-4018    

DOI:10.1139/o77-007

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

Pyruvate kinase (EC 2.7.1.40) ofNeurospora, a tetramer composed of apparently identical subunits, has been shown to be a dimer of dimers by interprotomeric cross-linking experiments in which bifunctional reagents were used. An analysis of the polyacrylamide gel profiles of the enzyme after cross-linking with glutaraldehyde, dimethyl suberimidate, and dimethyl adipimidate shows that the extent of intersubunit cross-linking is influenced markedly by the ligand bound to the enzyme. Bifunctional cross-linking reagents with a shorter distance between the two functional groups form cross-links effectively in the unliganded enzyme. In the FDP – pyruvate kinase complex, cross-linking was observed over longer distances compared with the unliganded enzyme. It is demonstrated that covalent cross-linkers can be used as sensitive indicators of conformational changes induced in pyruvate kinase by substrates and allosteric ligands.

ISSN:0008-4018    

DOI:10.1139/o77-008

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

The phytoestrogen formononetin was injected intramuscularly as [4-14C]formononetin into two adult hens. Radioactive materials in the urine for the succeeding 14 days (hen 1) or 16 days (hen 2) were fractionated on DEAE-Sephadex-25 columns by elution with a gradient of NaCl; the four major fractions thus separated were examined by solvent partition, thin-layer chromatography, and enzymic cleavage. The following seven radioactive components were identified in the urine, the average proportions of each being given in terms of percentage of total14C recovered from the urine: [14C]formononetin (4.3%); [14C]daidzein (11.4%); [14C]equol (6.8%); [14C]daidzein monosulfate (30.4%); [14C]equol monosulfate (5.8%); [14C]daidzein disulfate (19.8%); and [14C]equol disulfate (6.5%). Small proportions of sulfates of unidentified radioactive phenols were present. Tests for presence of glucosiduronates of14C-labelled material gave negative results. Radioactive formononetin sulfate was not detected in the urine of either hen.

ISSN:0008-4018    

DOI:10.1139/o77-009

出版商:NRC Research Press    

年代:1977

数据来源: NRC

摘要:

The effects of a series of inhibitors of monoamine oxidase (EC 1.4.3.4) and diamine oxidase (EC 1.4.3.6) and of two chelating agents were studied in rats, with respect to the catabolism of labeled pentylamine and putrescine to radioactive carbon dioxide,D-Tranylcypromine, clorgyline, and deprenyl inhibited oxidation of the monoamine, with essentially no effect on putrescine, under our test conditions. Aminoguanidine inhibited putrescine but not pentylamine oxidation. Iproniazid, isoniazid, and Lilly 51641 affected the catabolism of both amines. Pargyline inhibited putrescine oxidation, apparently in a reversible manner.

ISSN:0008-4018    

DOI:10.1139/o77-010

出版商:NRC Research Press    

年代:1977

数据来源: NRC