PubMed Health⌕ Search

Biomedical subjects

J Millet

Publications and source records attributed to J Millet.

17 recordsLinked to original sources
PubMed1992

Cellulose degradation by Clostridium thermocellum: from manure to molecular biology.

Clostridium thermocellum, a Gram-positive, thermophilic anaerobe produces a highly active cellulase system. This system, termed the cellulosome, is a complex composed of at least 14-18 different types of components organized around a large, cellulose-binding protein. Combining recombinant DNA technology and protein biochemistry has proved to be a successful approach in unravelling some important features of the system.

Amino Acid Sequence↗
PubMed1992

The venous antithrombotic effect of LF 1351 in the rat following oral administration.

The venous antithrombotic effects of a novel chemical entity, LF 1351, were investigated in rats following single oral administration, in comparison with i.v. administered heparin. LF 1351 demonstrated a dose-related antithrombotic effect in three models of venous thrombosis. The compound was approximately equipotent in two models involving complete stasis of the vena cava and administration of factor Xa or porcine serum, giving respective ED50 values of 48.7 mg/kg and 36.7 mg/kg. LF 1351 was less effective in a model involving partial stasis in the presence of an endothelial lesion. In this case, the antithrombotic effect did not exceed 60-65%, the ED50 being 150 mg/kg. Heparin (50-300 micrograms/kg; 7.5-45.0 U/kg) was effective in all three models. At the approximate ED80 value against factor Xa-induced thrombosis (75 mg/kg) the antithrombotic effect of LF 1351 persisted for 6 h. The antithrombotic effect of LF 1351 (300 mg/kg) occurred without significant changes in APTT or thrombin time.

Administration, Oral↗
PubMed1991

Identification of a histidyl residue in the active center of endoglucanase D from Clostridium thermocellum.

Diethylpyrocarbonate modification of endoglucanase D from Clostridium thermocellum, cloned in Escherichia coli, resulted in a rapid but partial (maximally 70-80%) loss of activity. The second-order rate constant of inactivation proved to be exceptionally high (3210 M-1.min-1). A 3-fold reduction of the kcat and a 2-fold increase of the Km for 2'-chloro-4'-nitrophenyl beta-cellobioside were observed. Spectrophotometric analysis indicate the presence of one rapidly (k = 0.45 min-1) and two slower (k = 0.23 min-1) reacting histidyl residues. In the presence of 50 mM methyl beta-cellotrioside, the rate of inactivation was reduced 16-fold, and the kinetics of modification were compatible with the protection of 1 histidyl residue. Since peptide analysis was inconclusive, identification of the critical residue was attempted by site-directed mutagenesis. Each of the 12 histidyl residues present in the endoglucanase D sequence was mutated into either Ala or Ser. Seven of the mutant enzymes had specific activities lower than 50% of the wild-type. Only in the case of the Ser-516 mutant, however, was the residual activity not affected by diethyl pyrocarbonate. These findings suggest an important functional or structural role for His-516 in the wild-type enzyme.

Amino Acid Sequence↗
PubMed1991

High activity of inclusion bodies formed in Escherichia coli overproducing Clostridium thermocellum endoglucanase D.

The formation of cytoplasmic inclusion bodies by Escherichia coli overproducing Clostridium thermocellum endoglucanase D (EGD) was investigated. EGD was found in inclusion bodies as a 68 kDa form, whereas the size of the cytoplasmic form was 65 kDa. Upon solubilization with urea followed by dialysis, the 68 kDa form was converted to the 65 kDa species. Proteolysis occurred within the COOH-terminal, reiterated region of the 68 kDa form, which is conserved among most C. thermocellum endoglucanases, but is not required for catalytic activity. The specific activity of the enzyme embedded in inclusion bodies was close to that of the purified protein. Thus, inclusion body formation does not involve denaturation of the catalytic domain of EGD, but, more likely, the participation of the reiterated, conserved region in intermolecular interactions.

Blotting, Western↗
PubMed1991

Peak blistering point: influence on fluid levels of 5-MOP in human skin in vivo after systemic administration.

The concentration of 5-methoxypsoralen (5-MOP) in suction blister fluid (SBF) after oral intake was determined in relation to the peak blistering point. Interstitial fluid was obtained from nine healthy male volunteers by applying mild suction (300-350 mmHg) to the skin of the volar aspect of the forearm. Blisters were raised at three different times: 18 h prior to drug administration (group I); 2 h prior to drug administration (group II); and during drug ingestion (group III). SBF levels of 5-MOP were determined in each group of blisters 2 h after oral administration of 1.2 mg/kg micronized 5-MOP. The results showed a statistically significant difference in the concentrations of 5-MOP between each group of blisters. The highest concentration of 5-MOP in the SBF was found in group III and the lowest in group II blisters. These findings suggest that the point of peak blistering in relation to drug administration needs to be known in each pharmacological study using suction blisters.

5-Methoxypsoralen↗
PubMed1991

[Galenic forms].

Explore the source record for details and available documents.

Administration, Inhalation↗
PubMed1989

Evaluation of the effect of heparin and tetracycline on the cohesion of the dermal-epidermal junction.

In an attempt to demonstrate the effect of tetracycline or heparin therapy on the dermo-epidermal junction, the suction blister time was measured at the beginning and at the end of the study in 18 female Sprague-Dawley hairless rats, using the suction blister method. Suction blister time was defined as the time taken for the first sign of a vesicle to appear in the hole in the diaphragm of a suction chamber. 200 IU heparinate calcium was administered daily for 6 days in 6 rats. Six other rats received 27.5 mg tetracycline chlorhydrate daily for 6 days. The others did not receive any drug. Significant statistical differences in suction blister time before and after administration of the drugs were found in the heparin group (p less than 0.05), and in the tetracycline group (p less than 0.05), compared with the control group. Heparin and tetracycline were found to increase suction blister time significantly. These results suggest an increased cohesion in the dermo-epidermal junction due to heparin and tetracycline.

Animals↗
PubMed1986

Purification and properties of the endoglucanase C of Clostridium thermocellum produced in Escherichia coli.

The celC gene, which codes for a new endoglucanase of Clostridium thermocellum, termed endoglucanase C, was found to be expressed when cloned in Escherichia coli. The enzyme was purified to electrophoretic homogeneneity from E. coli and its biochemical properties were studied. It differs from the previously studied endoglucanases A and B. In particular, endoglucanase C displays features common to endo- and exoglucanases, since it had a high activity on carboxymethylcellulose and on p-nitrophenyl-beta-D-cellobioside where only the agluconic bond was split. In addition, the enzyme was able to release cellobiose units from G3, G4 and G5 cellodextrins. Endoglucanase C was characterized by Western blot in a culture supernatant from C. thermocellum grown on cellulose, using an antiserum raised against the enzyme produced by E. coli.

Carboxymethylcellulose Sodium↗
PubMed1979

Characterization of an inhibitor of the intracellular protease from Bacillus subtilis.

A specific inhibitor of intracellular serylprotease from Bacillus subtilis has been isolated from both growing and sporulating cells. Like other protease inhibitors isolated from eukaryotic cells, the inhibitor from B. subtilis is a thermostable protein. A purification method is described. The molecular weight estimated by Biogel filtration and SDS gel electrophoresis is about 15,500. Both proteolytic and esterolytic activities of intracellular protease are equally sensitive to inhibition. With azocoll or Z-tyrosine p-nitrophenylester as substrates, noncompetitive inhibition patterns are observed. The inhibitor has no effect on the proteolytic or esterolytic activities of the extracellular serylprotease. A similar thermostable inhibitor is also present in Bacillus megaterium.

Bacillus megaterium↗
PubMed1976

[Thermosensitive sporulation and extracellular serylprotease mutant of B. subtilis].

Isolation and properties of B. subtilis ts 19 mutant, isolated as thermosensitive for sporulation, are described. At the non permissive temperature (42degreesC), the mutant cells are blocked at stage zero of sporulation and do not excrete extracellular enzymes such as serylprotease and esterase. At the permissive temperature (30degreesC), sporulation and excretion of extracellular enzymes are normal but the serylprotease is modified in its structure. Two molecular forms of this enzyme can be separated by polyacrylamide elecctrophoresis, both more thermolabile than the corresponding enzyme of the mother strain. Experiments of reversion and of transformation for the sporulation character have suggested that ts 19 contained two independent thermosensitive mutations. One of them is responsible for the pleiotropic Spo OA phenotype at the non permissive temperature. The other mutation is likely to reside in the structural gene coding for the extracellular serylprotease and leads to the formation of a modified enzyme which hydrolyzes itself into at least two types of more stable molecules. No conclusion can be drawn with certainty concerning the physiologi-al role of the extracellular serylprotease in sporulation. It may be pointed out however that transformants for the Sp+ character at 42degreesC keep the same impaired serylprotease as the ts 19 mutant and sporulate, at any temperature, as well as the wild strain.

Bacillus subtilis↗
PubMed1975

[Purification and properties of an esterase excreted during sporulation of Bacillus subtilis].

After growth in Difco Nutrient Broth, several proteolytic enzymes are excreted by B. subtilis, Marburg strain, namely a metalloprotease and a seryl protease. We report here the purification and some biochemical properties of a third extracellular hydrolytic enzyme for which we propose the term of esterase. As the serylprotease, the esterase is a seryl enzyme endowed with both proteolytic and esterolytic activities. Nevertheless the esterase differs from serylprotease in many aspects. In particular, it is an acidic enzyme with a low proteolytic activity and a high esterolytic activity. Its specificity toward synthetic substrates is restricted. The enzyme is active only on esters of amino acids and particularly on those of tyrosine. With Bz Tyr O Et as substrate the esterase displays a maximum of activity between pH 7.2 and 8.1 with a Km of 1.3.10-minus 3 M at 30 degrees C. At the end of this work, two questions remain unanswered: 1) the origin of the multiplicity of the bands revealed by polyacrylamide electrophoresis in the purest fraction; 2) the nature of the physiological substrate of the enzyme.

2,2'-Dipyridyl↗
PubMed1975

[Tablets. XII].

Explore the source record for details and available documents.

Pressure↗