James De La Rosa | Pima Community College (original) (raw)
Uploads
Papers by James De La Rosa
The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from... more The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from fed rats and cats. In incubations of rat hepatocytes with cysteinesulfinate, the rate of hypotaurine plus taurine production was approximately the same as the rate of conversion of the 1-carbon of cysteinesulfinate to CO.,. In contrast, no significant production of hypotaurine plus taurine occurred in incubations of
Bioluminescence and Chemiluminescence, 1981
Bioluminescence and Chemiluminescence, 1981
Comparative biochemistry and physiology. B, Comparative biochemistry, 1985
The relationship between activities of enzymes involved in cysteine oxidation and the apparent co... more The relationship between activities of enzymes involved in cysteine oxidation and the apparent conversion of cysteine to taurine in vivo were investigated in the rat and cat. Both hepatic cysteinesulfinate decarboxylase activity and the oxidation in vivo of cysteine to taurine were lower in the kitten than in the adult female rat and lower in the latter than in the young male rat. Our data support the hypothesis that cysteinesulfinate decarboxylase plays a rate-limiting role in taurine biosynthesis.
The Journal of nutrition, 1987
The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from... more The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from fed rats and cats. In incubations of rat hepatocytes with cysteinesulfinate, the rate of hypotaurine plus taurine production was approximately the same as the rate of conversion of the 1-carbon of cysteinesulfinate to CO2. In contrast, no significant production of hypotaurine plus taurine occurred in incubations of cat hepatocytes with cysteinesulfinate. These data are consistent with the species difference in the activity of hepatic cysteinesulfinate decarboxylase, which converts cysteinesulfinate to hypotaurine. In incubations of either rat or cat hepatocytes with cysteine, no hypotaurine plus taurine production was detected. However, the 1-carbon of cysteine was converted to CO2 and the production of urea plus ammonia nitrogen was significantly increased over the rates observed in incubations of cells without substrate. Our results suggest that most cysteine oxidation by hepatocytes ...
Journal of chromatography, Jan 11, 1985
Cysteinesulfinate, hypotaurine and taurine, which are key metabolites of cysteine, can be separat... more Cysteinesulfinate, hypotaurine and taurine, which are key metabolites of cysteine, can be separated from each other and other closely eluting amino acids in biological samples by reversed-phase high-performance liquid chromatography on a Waters Nova-Pak C18 column. Samples were derivatized with o-phthalaldehyde-2-mercaptoethanol prior to injection. The elution system consisted of 100 mM potassium phosphate buffer, pH 7.0, with 3% (v/v) tetrahydrofuran with an initial isocratic phase at 1.2% acetonitrile and a gradient from 1.2 to 12.8% acetonitrile. This method is suitable for measurement of the production of metabolites from cysteine by isolated cells and for analysis of plasma and tissue extracts. Low levels of hypotaurine in rat tissues were easily measured with this method and are reported here for the first time.
The Journal of nutrition, 1990
Both cysteinesulfinate-independent and cysteinesulfinate-dependent pathways are involved in the c... more Both cysteinesulfinate-independent and cysteinesulfinate-dependent pathways are involved in the catabolism of cyst(e)ine by freshly isolated rat renal cortical tubules. Sulfate and thiosulfate were shown to be the major sulfur-containing products that accumulated in incubations of renal tubules with 1 mmol/L or 25 mmol/L [35S]cyst(e)ine. Thiosulfate is an intermediate in the oxidation of the sulfide produced by the cysteinesulfinate-independent catabolism of cyst(e)ine by desulfhydration pathway(s), whereas sulfate is formed both by further oxidation of thiosulfate and by oxidation of the sulfite formed by the cysteinesulfinate-transamination pathway. Incubation of renal tubules with propargylglycine inhibited gamma-cystathionase activity by 85%, and this resulted in a 46% decrease in sulfate production and a 68% decrease in thiosulfate production when the treated renal tubules were incubated with 1 mmol/L [35S]cyst(e)ine. Addition of 25 mmol/L unlabeled cysteinesulfinate to create ...
Cancer research, Jan 15, 1992
We have recently reported that methionine adenosyltransferase (MAT) in resting human peripheral b... more We have recently reported that methionine adenosyltransferase (MAT) in resting human peripheral blood T-cells is primarily present in the form of a precursor which we named lambda. This protein decreases upon cell stimulation, as both MAT activity and the amount of the catalytic alpha/alpha' subunits of the enzyme increase. When resting cells are activated by phytohemagglutinin, the decrease in lambda and increase in alpha/alpha' occurs after interleukin 2 (IL-2) production and before DNA synthesis. The human T-leukemia cell line, Jurkat, is unique in its ability to produce IL-2 in response to exogenous stimuli such as T-cell mitogens and therefore provides a convenient model for studying biochemical reactions involved in T-cell activation. In this study the regulation of MAT activity and S-adenosylmethionine (AdoMet) in resting and activated Jurkat cells was investigated. Here we report that MAT activity in unstimulated Jurkat cells is about 10- and 3-fold higher than the a...
Methods in Enzymology, 1987
Life Sciences, 1985
~dministration of guanidinoethanesulfonate (GES) to male rats for 5 weeks resulted in a 90% decre... more ~dministration of guanidinoethanesulfonate (GES) to male rats for 5 weeks resulted in a 90% decrease in the hepatic taurine concentration. This depletion of hepatic taurine was associated with a 570% increase in the concentration of glycine-conjugated bile acids, a 30 % decrease in the concentration of taurine-conjugated bile acids, and an increase in the ratio of glycine-to taurine-conjugated bile acids from 0.046 to 0.45. The total concentration of bile salts in the bile and the turnover of cholic acid were not affected by administration of GE~ The data indicate that the taurine-depleted rat conserves taurine to some extent by using glycine instead of taurine for bile salt synthesis but not by decreasing the daily fractional turnover of bile acids.
Journal of Biological Chemistry, 1995
Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 1991
S-Adenosylmethionine (AdoMet), inorganic pyrophosphate (PPi) and inorganic phosphate (Pi) are pot... more S-Adenosylmethionine (AdoMet), inorganic pyrophosphate (PPi) and inorganic phosphate (Pi) are potent product inhibitors of AdoMet synthetase and have been postulated to play a role in increasing AdoMet levels and turnover in peripheral blood mononuclear cells (PBM) after stimulation with phytohemagglutinin (PHA). Measurements of these metabolites in PHA-stimulated PBM showed the expected 2- to 3-fold increases in AdoMet after 8 h, and smaller increases in PPi and Pi. Since the kinetic model requires substantial decreases in PPi and Pi in response to PHA, product inhibition cannot explain the observed changes in AdoMet metabolism in this system. A 2.5-fold increase in AdoMet synthetase catalytic activity was found in crude extracts of PBM within 8 h of PHA-stimulation and probably accounts for increased cellular levels and utilization of AdoMet. Immunochemical analyses with a monoclonal antibody specific for the alpha/alpha' subunits of human lymphocyte AdoMet synthetase showed that these increases in catalytic activity were not associated with increases in immunoreactive protein. The ratio of catalytic activity to immunoreactivity in stimulated cells was 4-fold higher than in unstimulated controls and almost identical to that found in extracts from the human B-lymphocyte line WI-L2. Unstimulated PBM appear to contain substantial amounts of AdoMet synthetase alpha/alpha' subunit with reduced or absent catalytic activity, which can be activated by PHA-stimulation.
Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 1990
Although the physical and kinetic properties of S-adenosylmethionine (AdoMet) synthetases from di... more Although the physical and kinetic properties of S-adenosylmethionine (AdoMet) synthetases from different sources are quite different, it appears that these enzymes have structurally or antigenically conserved regions as demonstrated by studies with AdoMet synthetase specific antibodies. Polyclonal anti-human lymphocyte AdoMet synthetase crossreacted with enzyme from rat liver (fl isozyme), Escherichia coil and yeast. In addition, polycional anti-E, coli enzyme and antibodies to synthetic peptides copying several regions of the yeast enzyme reacted with the human V and rat fl isozymes. Antibodies to yeast SAM1 encoded protein residues 6-21, 87-113 and 87-124 inhibited the activity of human lymphocyte AdoMet synthetase, while antibodies to residues 272-287 had no effect on the enzyme activity. Our results suggest that these conserved regions may be important in enzyme activity.
Biochimica et Biophysica Acta (BBA) - General Subjects, 1994
Two peaks of methionine adenosyltransferase (MAT) activity from human erythrocytes were partially... more Two peaks of methionine adenosyltransferase (MAT) activity from human erythrocytes were partially purified on a DEAE-cellulose column. Using anti-MAT antibodies, a 60 kDa form of MAT, referred to as rho was identified in peak I. Although rho represented the major MAT protein in crude erythrocyte extracts, the enzyme was very labile and accounted for only 6% of the total MAT activity. Peak II enzyme was stable, and consisted of the previously described catalytic alpha (53 kDa) subunit and the beta subunit (38 kDa), both of which are found in activated human lymphocytes and leukemic cells of lymphoid origin. Mature normal and polycythemic erythrocytes contained predominantly rho as the major MAT protein, while nucleated erythrocytes and reticulocytes contained predominantly the lambda (68 kDa), the major form found in resting human lymphocytes. Human erythroleukemic cells (HEL 92.1.7) contained the alpha, alpha' and beta subunits of MAT, and in this regard was indistinguishable from MAT found in activated lymphocytes and leukemic cells of lymphoid origin (Jurkat). Since rho was generated during the incubation of extracts from resting lymphocytes, which contain predominantly lambda, in the absence of protease inhibitors; the rho form of MAT appears to be derived from the lambda form by proteolytic cleavage. The data indicate that distinct forms of MAT are present at different stages of erythrocyte maturation and reveal the presence of a new form of MAT with reduced activity compared to previously described forms.
The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from... more The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from fed rats and cats. In incubations of rat hepatocytes with cysteinesulfinate, the rate of hypotaurine plus taurine production was approximately the same as the rate of conversion of the 1-carbon of cysteinesulfinate to CO.,. In contrast, no significant production of hypotaurine plus taurine occurred in incubations of
Bioluminescence and Chemiluminescence, 1981
Bioluminescence and Chemiluminescence, 1981
Comparative biochemistry and physiology. B, Comparative biochemistry, 1985
The relationship between activities of enzymes involved in cysteine oxidation and the apparent co... more The relationship between activities of enzymes involved in cysteine oxidation and the apparent conversion of cysteine to taurine in vivo were investigated in the rat and cat. Both hepatic cysteinesulfinate decarboxylase activity and the oxidation in vivo of cysteine to taurine were lower in the kitten than in the adult female rat and lower in the latter than in the young male rat. Our data support the hypothesis that cysteinesulfinate decarboxylase plays a rate-limiting role in taurine biosynthesis.
The Journal of nutrition, 1987
The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from... more The metabolism of cysteine and cysteinesulfinate was studied in freshly isolated hepatocytes from fed rats and cats. In incubations of rat hepatocytes with cysteinesulfinate, the rate of hypotaurine plus taurine production was approximately the same as the rate of conversion of the 1-carbon of cysteinesulfinate to CO2. In contrast, no significant production of hypotaurine plus taurine occurred in incubations of cat hepatocytes with cysteinesulfinate. These data are consistent with the species difference in the activity of hepatic cysteinesulfinate decarboxylase, which converts cysteinesulfinate to hypotaurine. In incubations of either rat or cat hepatocytes with cysteine, no hypotaurine plus taurine production was detected. However, the 1-carbon of cysteine was converted to CO2 and the production of urea plus ammonia nitrogen was significantly increased over the rates observed in incubations of cells without substrate. Our results suggest that most cysteine oxidation by hepatocytes ...
Journal of chromatography, Jan 11, 1985
Cysteinesulfinate, hypotaurine and taurine, which are key metabolites of cysteine, can be separat... more Cysteinesulfinate, hypotaurine and taurine, which are key metabolites of cysteine, can be separated from each other and other closely eluting amino acids in biological samples by reversed-phase high-performance liquid chromatography on a Waters Nova-Pak C18 column. Samples were derivatized with o-phthalaldehyde-2-mercaptoethanol prior to injection. The elution system consisted of 100 mM potassium phosphate buffer, pH 7.0, with 3% (v/v) tetrahydrofuran with an initial isocratic phase at 1.2% acetonitrile and a gradient from 1.2 to 12.8% acetonitrile. This method is suitable for measurement of the production of metabolites from cysteine by isolated cells and for analysis of plasma and tissue extracts. Low levels of hypotaurine in rat tissues were easily measured with this method and are reported here for the first time.
The Journal of nutrition, 1990
Both cysteinesulfinate-independent and cysteinesulfinate-dependent pathways are involved in the c... more Both cysteinesulfinate-independent and cysteinesulfinate-dependent pathways are involved in the catabolism of cyst(e)ine by freshly isolated rat renal cortical tubules. Sulfate and thiosulfate were shown to be the major sulfur-containing products that accumulated in incubations of renal tubules with 1 mmol/L or 25 mmol/L [35S]cyst(e)ine. Thiosulfate is an intermediate in the oxidation of the sulfide produced by the cysteinesulfinate-independent catabolism of cyst(e)ine by desulfhydration pathway(s), whereas sulfate is formed both by further oxidation of thiosulfate and by oxidation of the sulfite formed by the cysteinesulfinate-transamination pathway. Incubation of renal tubules with propargylglycine inhibited gamma-cystathionase activity by 85%, and this resulted in a 46% decrease in sulfate production and a 68% decrease in thiosulfate production when the treated renal tubules were incubated with 1 mmol/L [35S]cyst(e)ine. Addition of 25 mmol/L unlabeled cysteinesulfinate to create ...
Cancer research, Jan 15, 1992
We have recently reported that methionine adenosyltransferase (MAT) in resting human peripheral b... more We have recently reported that methionine adenosyltransferase (MAT) in resting human peripheral blood T-cells is primarily present in the form of a precursor which we named lambda. This protein decreases upon cell stimulation, as both MAT activity and the amount of the catalytic alpha/alpha' subunits of the enzyme increase. When resting cells are activated by phytohemagglutinin, the decrease in lambda and increase in alpha/alpha' occurs after interleukin 2 (IL-2) production and before DNA synthesis. The human T-leukemia cell line, Jurkat, is unique in its ability to produce IL-2 in response to exogenous stimuli such as T-cell mitogens and therefore provides a convenient model for studying biochemical reactions involved in T-cell activation. In this study the regulation of MAT activity and S-adenosylmethionine (AdoMet) in resting and activated Jurkat cells was investigated. Here we report that MAT activity in unstimulated Jurkat cells is about 10- and 3-fold higher than the a...
Methods in Enzymology, 1987
Life Sciences, 1985
~dministration of guanidinoethanesulfonate (GES) to male rats for 5 weeks resulted in a 90% decre... more ~dministration of guanidinoethanesulfonate (GES) to male rats for 5 weeks resulted in a 90% decrease in the hepatic taurine concentration. This depletion of hepatic taurine was associated with a 570% increase in the concentration of glycine-conjugated bile acids, a 30 % decrease in the concentration of taurine-conjugated bile acids, and an increase in the ratio of glycine-to taurine-conjugated bile acids from 0.046 to 0.45. The total concentration of bile salts in the bile and the turnover of cholic acid were not affected by administration of GE~ The data indicate that the taurine-depleted rat conserves taurine to some extent by using glycine instead of taurine for bile salt synthesis but not by decreasing the daily fractional turnover of bile acids.
Journal of Biological Chemistry, 1995
Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 1991
S-Adenosylmethionine (AdoMet), inorganic pyrophosphate (PPi) and inorganic phosphate (Pi) are pot... more S-Adenosylmethionine (AdoMet), inorganic pyrophosphate (PPi) and inorganic phosphate (Pi) are potent product inhibitors of AdoMet synthetase and have been postulated to play a role in increasing AdoMet levels and turnover in peripheral blood mononuclear cells (PBM) after stimulation with phytohemagglutinin (PHA). Measurements of these metabolites in PHA-stimulated PBM showed the expected 2- to 3-fold increases in AdoMet after 8 h, and smaller increases in PPi and Pi. Since the kinetic model requires substantial decreases in PPi and Pi in response to PHA, product inhibition cannot explain the observed changes in AdoMet metabolism in this system. A 2.5-fold increase in AdoMet synthetase catalytic activity was found in crude extracts of PBM within 8 h of PHA-stimulation and probably accounts for increased cellular levels and utilization of AdoMet. Immunochemical analyses with a monoclonal antibody specific for the alpha/alpha' subunits of human lymphocyte AdoMet synthetase showed that these increases in catalytic activity were not associated with increases in immunoreactive protein. The ratio of catalytic activity to immunoreactivity in stimulated cells was 4-fold higher than in unstimulated controls and almost identical to that found in extracts from the human B-lymphocyte line WI-L2. Unstimulated PBM appear to contain substantial amounts of AdoMet synthetase alpha/alpha' subunit with reduced or absent catalytic activity, which can be activated by PHA-stimulation.
Biochimica et Biophysica Acta (BBA) - Protein Structure and Molecular Enzymology, 1990
Although the physical and kinetic properties of S-adenosylmethionine (AdoMet) synthetases from di... more Although the physical and kinetic properties of S-adenosylmethionine (AdoMet) synthetases from different sources are quite different, it appears that these enzymes have structurally or antigenically conserved regions as demonstrated by studies with AdoMet synthetase specific antibodies. Polyclonal anti-human lymphocyte AdoMet synthetase crossreacted with enzyme from rat liver (fl isozyme), Escherichia coil and yeast. In addition, polycional anti-E, coli enzyme and antibodies to synthetic peptides copying several regions of the yeast enzyme reacted with the human V and rat fl isozymes. Antibodies to yeast SAM1 encoded protein residues 6-21, 87-113 and 87-124 inhibited the activity of human lymphocyte AdoMet synthetase, while antibodies to residues 272-287 had no effect on the enzyme activity. Our results suggest that these conserved regions may be important in enzyme activity.
Biochimica et Biophysica Acta (BBA) - General Subjects, 1994
Two peaks of methionine adenosyltransferase (MAT) activity from human erythrocytes were partially... more Two peaks of methionine adenosyltransferase (MAT) activity from human erythrocytes were partially purified on a DEAE-cellulose column. Using anti-MAT antibodies, a 60 kDa form of MAT, referred to as rho was identified in peak I. Although rho represented the major MAT protein in crude erythrocyte extracts, the enzyme was very labile and accounted for only 6% of the total MAT activity. Peak II enzyme was stable, and consisted of the previously described catalytic alpha (53 kDa) subunit and the beta subunit (38 kDa), both of which are found in activated human lymphocytes and leukemic cells of lymphoid origin. Mature normal and polycythemic erythrocytes contained predominantly rho as the major MAT protein, while nucleated erythrocytes and reticulocytes contained predominantly the lambda (68 kDa), the major form found in resting human lymphocytes. Human erythroleukemic cells (HEL 92.1.7) contained the alpha, alpha' and beta subunits of MAT, and in this regard was indistinguishable from MAT found in activated lymphocytes and leukemic cells of lymphoid origin (Jurkat). Since rho was generated during the incubation of extracts from resting lymphocytes, which contain predominantly lambda, in the absence of protease inhibitors; the rho form of MAT appears to be derived from the lambda form by proteolytic cleavage. The data indicate that distinct forms of MAT are present at different stages of erythrocyte maturation and reveal the presence of a new form of MAT with reduced activity compared to previously described forms.