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C Rojas

Publications and source records attributed to C Rojas.

At least 19 recordsLinked to original sources
PubMed1992

Two DNA polymerases from Trypanosoma cruzi: biochemical characterization and effects of inhibitors.

1. Two DNA polymerases have been partially purified from Trypanosoma cruzi epimastigotes by DEAE-cellulose, phosphocellulose, and DNA agarose chromatography. 2. Both enzyme activities were characterized by several biochemical criteria. 3. They showed different sensitivity to KCl and displayed characteristic Mg2+ and Mn2+ requirements, although they exhibited almost identical primer-template utilization. 4. The preferred substrates were poly dC-oligo dG, activated calf thymus DNA, and poly dT-oligo rA. 5. Both enzyme fractions are not inhibited by aphidicolin while N-ethylmaleimide and phosphonacetic acid inhibited them to different extents. 6. ButylphenyldGTP strongly inhibited T. cruzi enzyme fraction I while it had no effect on enzyme fraction III. 7. This dGTP analog also inhibited the poly dT-directed polymerization of dAMP as described for other mammalian DNA polymerases. Kinetic studies indicated that butylphenyldGTP inhibited enzyme fraction I in a non-competitive fashion.

Animals↗
PubMed1992

The genomic structure of the human skeletal muscle sodium channel gene.

Electrical excitability of neurons and muscle cells reflects the actions of a family of structurally related sodium channels. Mutations in the adult skeletal muscle sodium channel have been associated with the inherited neuromuscular disorders paramyotonia congenita (PMC) and hyperkalemic periodic paralysis (HPP). We have deciphered the entire genomic structure of the human skeletal muscle sodium channel gene and developed a restriction map of the locus. SCN4A consists of 24 exons spanning 35 kb of distance on chromosome 17q. We describe the sequence of all intron/exon boundaries, the presence of several polymorphisms in the coding sequence, and the locations within introns of two dinucleotide repeat polymorphisms. This is the first sodium channel for which the entire genomic structure has been resolved. The organization of the SCN4A exons relative to the proposed protein structure is presented and represents a foundation for functional and evolutionary comparisons of sodium channels. Knowledge of the exon structure and flanking intron sequences for SCN4A will permit a systematic search for mutations in PMC and HPP.

Amino Acid Sequence↗
PubMed1992

Sensitivity to in vitro lipid peroxidation in liver and brain of aged rats.

Lipid peroxidation in rat liver and brain has been studied to see if it increases with old age. No significant differences in the level of endogenous, nonstimulated lipid peroxidation (TBA-RS) is found between 9 month-old (mature adults) and 28 month-old animals in liver or cerebral cortex. Liver homogenates subjected in vitro to an oxidative stress (ascorbate-Fe++), show a clearly slower peroxidation rate in old than in young animals. On the other hand, the in vitro peroxidation rate of cerebral homogenates was similar in young and old animals. The in vitro peroxidation rate was much higher in brain than in liver tissue. These results do not support the view that old rats liver and brain are more susceptible to free radical oxidative damage than those of young ones.

Aging↗
PubMed1991

Simultaneous determination of two antioxidants, uric and ascorbic acid, in animal tissue by high-performance liquid chromatography.

A rapid and simple method for the simultaneous analysis of uric and ascorbic acid in extracts of animal tissue is described. The method uses reversed-phase ion-pair chromatography with ultraviolet detection. The technique allows efficient separation of both acids while showing high selectivity, recovery, reproducibility, and sample stability. Calculated levels of both substances in mouse liver tissue were 1.00 +/- 0.05 mumol ascorbic acid/g and 130 +/- 5 nmol uric acid/g.

Animals↗
PubMed1991

Comparison of the action of lipoprotein lipase on triacylglycerols and phospholipids when presented in mixed liposomes or in emulsion droplets.

We have compared the action of lipoprotein lipase on liposomes of egg yolk phosphatidylcholine containing less than saturating amounts of trioleoylglycerol (less than 3%) and emulsion droplets of the same lipids. The amounts of the two types of lipid particles (expressed in terms of phosphatidylcholine) needed to reach substrate saturation of the enzyme were similar, indicating similar binding of the lipase to these two lipid/water interfaces. With liposomes, as opposed to emulsion droplets, albumin was not necessary for continued hydrolysis of triacylglycerols, presumably because product fatty acids could be accommodated in the phospholipid bilayer. The maximal rate of trioleoylglycerol hydrolysis was more than 10-fold higher, and the ratio of trioleoylglycerol/phosphatidylcholine hydrolysis was more than 50-fold higher with the emulsion droplets. Qualitatively similar results were obtained with hepatic lipase, and a lipase from Pseudomonas fluorescence. The data suggest that the lipases remained at the interface for several catalytic cycles, and that a continued supply of substrate molecules to the active site favored triacylglycerol entry from the core of the lipid particle, rather than sliding in from the side through lateral diffusion in the surface layer.

Apolipoprotein C-II↗
PubMed1990

Synthesis and secretion of active lipoprotein lipase in Chinese-hamster ovary (CHO) cells.

Cultured Chinese-hamster ovary cells (CHO cells) were found to produce and secrete a lipase, which was identified as a lipoprotein lipase by the following criteria. Its activity was stimulated by serum and apolipoprotein CII, and was inhibited by high salt concentration. The lipase bound to heparin-agarose and co-eluted with 125I-labelled bovine lipoprotein lipase in a salt gradient. A chicken antiserum to bovine lipoprotein lipase inhibited the activity and precipitated a labelled protein of the same apparent size as bovine lipoprotein lipase from media of CHO cells labelled with [35S]methionine. The lipase activity and secretion were similar in growing cells and in cells that had reached confluency. Hence, lipoprotein lipase appears to be expressed constitutively in CHO cells and is not linked to certain growth conditions, as in pre-adipocyte and macrophage cell lines. At 37 degrees C, but not at 4 degrees C, heparin increased the release of lipase to the medium 2-4-fold. This increased release occurred without depletion of cell-associated lipase activity, suggesting that heparin enhanced release of newly synthesized lipase.

Animals↗
PubMed1990

Preliminary crystallographic study of the Erythrina rubrinervia lectin.

The galactose-specific lectin from Erythrina rubrinervia crystallizes in the hexagonal space group P6, (or P6(5)) with unit cell dimensions a = b = d = 135.1 A, c = 83.0 A. These parameters are compatible with the presence of a dimer with Mr = 60,000 in the asymmetric unit. The crystals are suitable for high-resolution X-ray analysis.

Crystallization↗
PubMed1990

Calcium release modulated by inositol trisphosphate in ruptured fibers from frog skeletal muscle.

To investigate the effect of inositol 1,4,5-trisphosphate on calcium release, we used fiber bundles of frog sartorius muscle mechanically permeabilized by a scratching procedure, and we detected increments in calcium concentration by measuring aqueorin light signals. Submicromolar concentrations of inositol 1,4,5-trisphosphate induced fast calcium-release signals, with a half time to peak of 60 ms or less. Similar responses were elicited by caffeine. The calcium-release signal induced by inositol 1,4,5-trisphosphate occurred at pCa values of 7 or lower, and the dose-response curve depended on the ionic composition of the incubation solution. Lower inositol 1,4,5-trisphosphate concentrations were needed to induce release when incubation solutions of ionic composition expected to depolarize the transverse tubule membrane were used. Inositol 1,4,5-trisphosphate was more effective than inositol 1,3,4-trisphosphate, inositol 1,4,5,6-tetrakisphosphate, and inositol 1,4-bisphosphate. The effect of inositol 1,4,5-trisphosphate was synergistic with that of caffeine, and was not inhibited by heparin. These results, by showing directly that at resting calcium levels inositol 1,4,5-trisphosphate elicited calcium release, are consistent with a role for inositol 1,4,5-trisphosphate as a chemical modulator in excitation/contraction coupling in skeletal muscle.

Aequorin↗
PubMed1990

A patient with features of both Bardet-Biedl and Alström syndromes.

We describe a 30-year-old patient with acanthosis nigricans, diabetes mellitus with insulin resistance, hypogonadotropic hypogonadism, pigmentary degeneration of the retina and cerebellar, pyramidal and posterior columnar involvement. He had normal mental function, normal hearing and no hexadactyly. The patient had symptoms of both Bardet-Biedl and Alström syndromes, but did not manifest all the main features of either syndrome. This suggests either that the Bardet-Biedl, Alström, Laurence-Moon syndromes (including the variant described by Edwards) have a highly variable expression or that our case is a new variant within this group of syndromes.

Acanthosis Nigricans↗
PubMed1990

The effects of cyclophosphamide on the gonadotrophic cells of the normal rat.

The effects of the administration of cyclophosphamide to male rats at doses of 400 mg/m2 for a period of 5 days and of 200 mg/m2 over two five-day cycles interrupted by a 21-day break reveal differences in the ultrastructural morphology of gonadotrophic cells between treated animals and normal controls. The ultrastructural data obtained coincide with the results obtained from the measurement of serum FSH and LH levels, indicating hypofunction. A significant fall was also recorded in blood testosterone levels. The two types of treatment did not give rise to morphological or functional differences.

Animals↗
PubMed1990

[Serologic response to a DNA recombinant vaccine against hepatitis B in natives of the Peruvian Amazonian jungle].

Large areas of the Amazon basin in Brazil, Colombia, Ecuador, and in the nonoriental region of the peruvian jungle have been found to be hyperendemic to Hepatitis B with high prevalence of asymptomatic carriers (11 to 25%) and, in more selected areas, Hepatitis Delta has been also reported. In the present report, we have studied 108 volunteers from six different Jivaroes communities living in a hyperendemic Hepatitis B area. They received 2 doses of DNA recombinant yeast derivated HBV vaccine. All the selected persons were HBsAb negatives, but many (80%) had antibodies to HBc. Following immunization schedule, 80% responded with the formation of HBsAb; a better seroconversion was achieved in those negatives to anticore IgG compared with those having HBcAb. We obtained 90% of seroconversion in spite of the fact that our vaccination schedule was prolonged up to 10 months from the one recommended by the manufacturer. The vaccination schedule 0,4, 14 months, and the schedule 0,4 months, had 76 and 29% of seroconversion, respectively. We want to point out three observations: 1) It is quite possible that many of the Anti-core positives, that did not respond to vaccination were carriers of HBsAg undetectable by the conventional EIA test carried out; 2) The seroconversion rate in these natives was low (up to six months after the vaccination schedule); and 3) Many of the HBcAb were false positives and many of them were recently infected. We conclude: A) It is highly important to assess the anti-HBs hyperendemic areas before attempting vaccinations; B) All persons negative to anti-HBs should be vaccinated in spite to anticore antibodies; C) Areas with difficult access could be vaccinated even until 10 months without affecting good results, and D) DNA recombinant vaccine (ENGERIX B) was well tolerated. No side effects were observed.

Adolescent↗
PubMed1989

Spectral observation of an acyl-enzyme intermediate of lipoprotein lipase.

There have been several studies indicating that hydrolysis reactions of fatty acid esters catalyzed by lipases proceed through an acyl-enzyme intermediate typical of serine proteases. In particular, one careful kinetic study with the physiologically important enzyme lipoprotein lipase (LPL) is consistent with rate-limiting deacylation of such an intermediate. To observe the spectrum of acyl-enzyme and study the mechanism of LPL-catalyzed hydrolysis of substrate, we have used a variety of furylacryloyl substrates including 1,2-dipalmitoyl-3-[(beta-2-furylacryloyl)triacyl]glyceride (DPFATG) to study the intermediates formed during the hydrolysis reaction catalyzed by the enzyme. After isolation and characterization of the molecular weight of adipose LPL, we determined its extinction coefficient at 280 nm to quantitate the formation of any acyl-enzyme intermediate formed during substrate hydrolysis. We observed an intermediate at low pH during the enzyme-catalyzed hydrolysis of (furylacryloyl)imidazole. This intermediate builds early in the reaction when a substantial amount of substrate has hydrolyzed but no product, furylacrylate, has been formed. The acyl-enzyme has a lambda max = 305 nm and a molar extinction coefficient of 22,600 M-1 cm-1; these parameters are similar to those for furylacryloyl esters including the serine ester. These data provide the first spectral evidence for a serine acyl-enzyme in lipase-catalyzed reactions. The LPL hydrolysis reaction is base catalyzed, exhibiting two pKa values; the more acidic of these is 6.5, consistent with base catalysis by histidine. The biphasic rates for substrate disappearance or product appearance and the absence of leaving group effect indicate that deacylation of intermediate is rate limiting.

Acylation↗
PubMed1989

The effects of cyclophosphamide on the prolactin cells of the normal rat.

Cyclophosphamide administered at doses of 400 mg/m2/5 days with sacrifice two days later, and 200 mg/m2/5 days with a 21-day break and a further five days of treatment, with sacrifice two days later, provokes similar effects. Functional activity is less marked in untreated and control animals than in treated ones, as the hormone data shows. But the ultrastructure of PRL cells in treated animals indicates the existence of clear hormone synthesis activity, evident in the fusion and clustering of granules at differing stages of maturity, etc.

Animals↗
PubMed1989

[Phase contrast microscopy in the detection of bacteriuria].

Despite the recognized superiority of traditional culture methods to investigate bacteriuria their routine use in population studies is costly. We proposed to use Phase contrast microscopy (PCM) which allows clear visualization of bacteria with no centrifugation or staining. Urine samples from 214 patients were studied by PCM and results compared to those obtained by quantitative culture. MCF had 95% sensitivity and 96% specificity. Therefore, PCM may be used to screen urine samples for further bacteriologic study with the highest chance of obtaining positive cultures.

Bacteriological Techniques↗
PubMed1988

Crystallization of the met repressor from Escherichia coli.

The met repressor from Escherichia coli has been crystallized in space group P21, with unit cell dimensions a = 35.6 A, b = 62.6 A, c = 44.5 A, beta = 102.4 degrees and one aporepressor dimer per asymmetric unit. Preliminary X-ray diffraction photographs show measurable intensities to beyond 1.5 A resolution, and the crystal form is ideally suited to high-resolution crystallographic analysis (1 A = 0.1 nm).

Amino Acid Sequence↗
PubMed1987

Preliminary crystallographic study of a complex between the Fab fragment of a monoclonal anti-lysozyme antibody (D1.3) and the Fab fragment from an anti-idiotopic antibody against D1.3.

An anti-lysozyme antibody, D1.3, was used as immunogen to obtain syngeneic (Balb/c) monoclonal anti-idiotopic antibodies. The complex between Fab D1.3 and the Fab fragment from the anti-idiotopic antibody E225 has been crystallized. The crystals are monoclinic, space group P2(1), with a = 75.7 A, b = 77.4 A, c = 97.2 A, beta = 111.90 degrees and one molecule of the complex in the asymmetric unit. X-ray photographs show reflections extending to a resolution of about 3 A. Although twinning occurs frequently in the large crystals obtained, this material is suitable for high-resolution X-ray analysis.

Antibodies, Monoclonal↗
PubMed1987

Schizodeme analyses of Trypanosoma cruzi zymodemes from Chile.

Kinetoplast DNA was isolated from Chilean Trypanosoma cruzi populations and digested with the restriction endonucleases EcoRI, HinfI, HpaII, MspI, and HaeIII. Three major schizodeme groups were discriminated. There was a correlation between the Chilean schizodeme groups (S1, S2, or S3) and the zymodemes known to occur in Chile (Z1, Brazilian Z2 and Bolivian Z2, respectively), although heterogeneity was seen within the schizodeme groups S2 and S3. Standard Brazilian and Bolivian T. cruzi clones (X10 clone 1, Esmeraldo clone 3, SC43 clone 1, and CAN III clone 1) and laboratory strains (Tulahuen and Y) were included in the schizodeme comparisons. SC43 clone 1 had obvious affinities with S3 and X10 clone 1 shared some features with S1 but the other reference stocks could not be definitely assigned to S1, S2, or S3. Fragment patterns and densitometric traces following digestion with HpaII or MspI suggested that kDNA sequences were not methylated.

Animals↗