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PubMed1990

Primary structure of a key enzyme in plant tetrapyrrole synthesis: glutamate 1-semialdehyde aminotransferase.

The formation of delta-aminolevulinate from glutamate 1-semialdehyde (GSA) is catalyzed by glutamate 1-semialdehyde aminotransferase (EC 5.4.3.8). The active form of the barley enzyme appears to be a dimer of identical subunits with a molecular mass of 46 kDa. From the purified enzyme, amino acid sequences of the N-terminal ends of the mature protein as well as an internal peptide were determined. DNA primers deduced from these peptide sequences were used to amplify with the polymerase chain reaction a cDNA sequence encoding part of the enzyme. Screening a cDNA library with this DNA fragment identified a full-length clone encoding the 49,540-Da precursor of the GSA aminotransferase. The transit peptide for chloroplast import consists of 34 amino acids. GSA aminotransferase and a precursor form were expressed on a multicopy plasmid in Escherichia coli. Both recombinant gene products reacted with an antibody against the barley GSA aminotransferase. Active barley GSA aminotransferase expressed in E. coli was shown to be active in assays of bacterial cell extracts. As a gene symbol for barley GSA aminotransferase, Gsa is proposed.

Amino Acid Sequence↗

PubMed2005

Determinants of substrate specificity in omega-aminotransferases.

Ornithine aminotransferase and 4-aminobutyrate aminotransferase are related pyridoxal phosphate-dependent enzymes having different substrate specificities. The atomic structures of these enzymes have shown (i) that active site differences are limited to the steric positions occupied by two tyrosine residues in ornithine aminotransferase and (ii) that, uniquely among related, structurally characterized aminotransferases, the conserved arginine that binds the alpha-carboxylate of alpha-amino acids interacts tightly with a glutamate residue. To determine the contribution of these residues to the specificities of the enzymes, we analyzed site-directed mutants of ornithine aminotransferase by rapid reaction kinetics, x-ray crystallography, and 13C NMR spectroscopy. Mutation of one tyrosine (Tyr-85) to isoleucine, as found in aminobutyrate aminotransferase, decreased the rate of the reaction of the enzyme with ornithine 1000-fold and increased that with 4-aminobutyrate 16-fold, indicating that Tyr-85 is a major determinant of specificity toward ornithine. Unexpectedly, the limiting rate of the second half of the reaction, conversion of ketoglutarate to glutamate, was greatly increased, although the kinetics of the reverse reaction were unaffected. A mutant in which the glutamate (Glu-235) that interacts with the conserved arginine was replaced by alanine retained its regiospecificity for the delta-amino group of ornithine, but the glutamate reaction was enhanced 650-fold, whereas only a 5-fold enhancement of the ketoglutarate reaction rate resulted. A model is proposed in which conversion of the enzyme to its pyridoxamine phosphate form disrupts the internal glutamate-arginine interaction, thus enabling ketoglutarate but not glutamate to be a good substrate.

Amino Acids↗

PubMed2003

Hepatic alanine-glyoxylate aminotransferase activity and oxalate metabolism in vitamin B6 deficient rats.

PURPOSE: Urinary oxalate has an important role in the formation of calcium oxalate stone and approximately 50% to 60% of urinary oxalate is derived from the endogenous metabolism of glyoxylate. Glyoxylate is enzymatically converted to glycine by alanine-glyoxylate aminotransferase in the liver and vitamin B6 has a key role as a coenzyme. Therefore, we evaluated hepatic alanine-glyoxylate aminotransferase activity and oxalate metabolism in vitamin B6 deficient rats. MATERIALS AND METHODS: A total of 12 male Wistar rats were fed a control or a vitamin B6 deficient diet. After 4 weeks creatinine, oxalate, glycolate, glycine and citrate in the urine, and alanine-glyoxylate aminotransferase activity and its mRNA level in the liver were measured. RESULTS: Urinary oxalate-to-creatinine and glycolate-to-creatinine ratios were significantly higher in vitamin B6 deficient rats than in control rats but urinary glycine-to-creatinine and citrate-to-creatinine ratios were significantly lower. Hepatic alanine-glyoxylate aminotransferase activity and its mRNA level were significantly lower in vitamin B6 deficient rats than in control rats. CONCLUSIONS: Vitamin B6 deficiency not only decreased alanine-glyoxylate aminotransferase activity, but also down-regulated alanine-glyoxylate aminotransferase gene expression by hepatocytes and led to hyperoxaluria and hyperglycolic aciduria secondary to impaired metabolism of oxalate precursors. Hyperoxaluria with hypocitruria may also contribute to calcium oxalate stone formation in vitamin B6 deficiency.

Animals↗

PubMed1978

Inhibition of glutamate:glyoxylate aminotransferase activity in tobacco leaves and callus by glycidate, an inhibitor of photorespiration.

The effect of glycidate (2,3-epoxypropionate), an inhibitor of glycolate synthesis and photorespiration in leaf tissue, was studied on glutamate:glyoxylate and serine:glyoxylate aminotransferases and glycine decarboxylase activities in particulate preparations obtained from tobacco (Nicotiana tabacum L.) callus and leaves. Glycidate specifically and effectively inhibited glutamate:glyoxylate aminotransferase. The inhibition was dependent on glycidate concentration and, to a lesser extent, on substrate concentration. The enzyme was not protected by either substrate. Even with saturating substrate concentrations the glycidate inhibition was only partially reversed. Under the in vitro assay conditions, glycidate inhibition of the aminotransferase was reversible. Glutamate:glyoxylate aminotransferase is the only enzyme of the glycolate pathway thus far examined which is severely inhibited by glycidate. However, in leaf discs, pretreatment with glycidate decreased both glutamate:glyoxylate and serine:glyoxylate aminotransferase activities suggesting binding by glycidate in vivo.Glycidate increased the pool sizes of both glutamate and glyoxylate in leaf discs. It has been shown that increases in concentration of either of these metabolites decrease photorespiration and glycolate synthesis and increase net photosynthesis. It is proposed that glycidate inhibits photorespiration indirectly by increasing the internal concentrations of glutamate and glyoxylate, as a consequence of the inhibition of glutamate:glyoxylate aminotransferase activity.

Journal Article↗

PubMed2006

Impaired health-related quality of life in patients with chronic hepatitis C and persistently normal aminotransferase levels.

A significant impact of hepatitis C virus (HCV) infection on health-related quality of life (HRQOL) has been previously described. However, comprehensive data on the quality of life in patients with chronic hepatitis C and persistently normal aminotransferase levels (PNAL) are currently not available. One hundred fifteen patients with chronic hepatitis C (45 with persistently normal aminotransferases and 70 with elevated aminotransferases) and 50 healthy subjects were enrolled. Emotional and psychological states were assessed by Profile of Mood States (POMS) scale and HRQOL was assessed by the 'Everyday Life' questionnaire (EDLQ), a validated questionnaire related to the SF-36 Health Survey. An impairment in HRQOL was observed in patients with chronic hepatitis C showing PNAL compared with healthy subjects with significant differences for the factor scores depression and anger in the POMS scale as well as for the items body, relationship to partner, self-confidence and zest of life in the EDLQ. No differences in any questionnaire were observed between patients with chronic hepatitis C showing PNAL or elevated aminotransferase levels except of a worse mean level for factor score anger in POMS scale in patients with persistently normal aminotransferases. No association of quality of life with severity of liver disease was found. Impairment of HRQOL by chronic infection with HCV is similar in patients with PNAL and those with elevated aminotransferase levels.

Adult↗

PubMed1985

Cloning and expression in Escherichia coli of cDNA for serine: pyruvate aminotransferase of rat liver.

Cloned cDNAs for rat liver serine: pyruvate aminotransferase were obtained by screening of a cDNA expression bank of rat liver with an antibody against the enzyme. Nineteen clones were isolated from 33 000 transformants and most of them had common fragments of cDNA on analysis by digestion with some restriction enzymes. These clones were identified as those containing cDNA for serine:pyruvate aminotransferase by the following criteria. (a) At the nucleic acid level, a 500-base-pair fragment of cDNA prepared by digestion of cDNAs with EcoRI and PstI hybridized with the mRNA coding for serine:pyruvate aminotransferase as judged by hybrid-selected and hybrid-arrested translations. (b) Specific proteins were detected in nine bacterial clones, a 40-kDa protein in one clone and a 39-kDa protein in eight clones. Among them only the 40-kDa protein was found to be solubilized from the cell by sonication, and this protein was immunoprecipitated with an antibody against serine:pyruvate aminotransferase of rat liver. (c) High activity of serine:pyruvate aminotransferase was expressed both in whole cell suspension and sonicated extract prepared from the transformant producing the 40-kDa protein, and 99% of the activity was immunoreactive with the antibody. Two types of mRNA for serine:pyruvate aminotransferase were detected on the RNA blot analysis by using cloned cDNA fragment as a probe. The larger mRNA (approximately 1600 nucleotides) was glucagon-inducible while the smaller one (approximately 1500 nucleotides) was not affected by the hormone.

Animals↗

PubMed1987

Nucleotide sequence of the cDNA encoding the precursor for mitochondrial serine:pyruvate aminotransferase of rat liver.

The nucleotide sequence of the mRNA coding for the precursor of mitochondrial serine:pyruvate aminotransferase of rat liver was determined from those of cDNA clones. The mRNA comprises at least 1533 nucleotides, except the poly(A) tail, and encodes a polypeptide consisting of 414 amino acid residues with a molecular mass of 45,834 Da. Comparison of the N-terminal amino acid sequence of mitochondrial serine:pyruvate aminotransferase with the nucleotide sequence of the mRNA showed that the mature form of the mitochondrial enzyme consisted of 390 amino acid residues of 43,210 Da. The amino acid composition of mitochondrial serine:pyruvate aminotransferase deduced from the nucleotide sequence of the cDNA showed good agreement with the composition determined on acid hydrolysis of the purified protein. The extra 24 amino acid residues correspond to the N-terminal extension peptide (pre-sequence) that is indispensable for the specific import of the precursor protein into mitochondria. In the extension peptide there are four basic amino acids distributed among hydrophobic amino acids and, as revealed on helical wheel analysis, the putative alpha-helical structure of the peptide was amphiphilic in nature. The secondary structures of the mature serine:pyruvate aminotransferase and three other aminotransferases of rat liver were predicted from their amino acid sequences. Their secondary structures exhibited a common feature and so we propose the specific lysine residue which binds pyridoxal phosphate as the active site of serine:pyruvate aminotransferase.

Amino Acids↗

PubMed1996

Cloning and expression of the gene for serine-glyoxylate aminotransferase from an obligate methylotroph Hyphomicrobium methylovorum GM2.

The gene encoding serine-glyoxylate aminotransferase, one of key enzymes for the assimilation of one-carbon compounds in methylotrophs, and its flanking regions were isolated from an obligate methylotrophic bacterium, Hyphomicrobium methylovorum GM2. Nucleotide sequencing of the recombinant plasmids revealed that the serine-glyoxylate aminotransferase gene encodes a 405-amino-acid protein with a calculated molecular mass of 43880 Da. The amino acid sequence of the enzyme showed identity to the sequences of serine-glyoxylate aminotransferase of Methylobacterium extorquens AM1 (57%), aspartate aminotransferase of Methanobacterium thermoformicicum (31%), human peroxisomal alanine-glyoxylate aminotransferase (27%), and serine-pyruvate aminotransferase of rat liver mitochondria (33%). The recombinant plasmid, which was constructed by ligation of the cloned gene and an expression vector pKK223-3, was introduced into Escherichia coli HB101. The recombinant enzyme was purified from transformed E. coli cells and analyzed by immunological and enzymological methods. The overexpressed enzyme was indistinguishable from the wild-type enzyme isolated from H. methylovorum GM2.

Amino Acid Sequence↗

PubMed2004

Branched-chain amino acid aminotransferase and methionine formation in Mycobacterium tuberculosis.

BACKGROUND: Tuberculosis remains a major world-wide health threat which demands the discovery and characterisation of new drug targets in order to develop future antimycobacterials. The regeneration of methionine consumed during polyamine biosynthesis is an important pathway present in many microorganisms. The final step of this pathway, the conversion of ketomethiobutyrate to methionine, can be performed by aspartate, tyrosine, or branched-chain amino acid aminotransferases depending on the particular species examined. RESULTS: The gene encoding for branched-chain amino acid aminotransferase in Mycobacterium tuberculosis H37Rv has been cloned, expressed, and characterised. The enzyme was found to be a member of the aminotransferase IIIa subfamily, and closely related to the corresponding aminotransferase in Bacillus subtilis, but not to that found in B. anthracis or B. cereus. The amino donor preference for the formation of methionine from ketomethiobutyrate was for isoleucine, leucine, valine, glutamate, and phenylalanine. The enzyme catalysed branched-chain amino acid and ketomethiobutyrate transamination with a Km of 1.77 - 7.44 mM and a Vmax of 2.17 - 5.70 micromol/min/mg protein, and transamination of ketoglutarate with a Km of 5.79 - 6.95 mM and a Vmax of 11.82 - 14.35 micromol/min/mg protein. Aminooxy compounds were examined as potential enzyme inhibitors, with O-benzylhydroxylamine, O-t-butylhydroxylamine, carboxymethoxylamine, and O-allylhydroxylamine yielding mixed-type inhibition with Ki values of 8.20 - 21.61 microM. These same compounds were examined as antimycobacterial agents against M. tuberculosis and a lower biohazard M. marinum model system, and were found to completely prevent cell growth. O-Allylhydroxylamine was the most effective growth inhibitor with an MIC of 78 microM against M. marinum and one of 156 microM against M. tuberculosis. CONCLUSION: Methionine formation from ketomethiobutyrate is catalysed by a branched-chain amino acid aminotransferase in M. tuberculosis. This enzyme can be inhibited by selected aminooxy compounds, which also have effectiveness in preventing cell growth in culture. These compounds represent a starting point for the synthesis of branched-chain aminotransferase inhibitors with higher activity and lower toxicity.

Amino Acid Sequence↗

PubMed1982

Purification and characterization of cysteine aminotransferase from rat liver cytosol.

Cysteine aminotransferase (L-cysteine: 2-oxoglutarate aminotransferase, EC 2.6.1.3) was purified over 400-fold from the high-speed supernatant fraction of rat liver. The purified enzyme was homogeneous as judged by gel filtration, isoelectric focusing and disc electrophoresis. The molecular weight of the enzyme was about 74,000 by gel filtration and the isoelectric point was 6.2 (4 degrees C). The enzyme catalyzed transamination between L-cysteine and 2-oxoglutarate and the reverse reaction. The optimum pH was 9.7. The Km value for L-cysteine was 22.2 mM, and that for 2-oxoglutaric acid was 0.06 mM. L-Aspartate was a potent inhibitor of the cysteine aminotransferase reaction. The enzyme was very active toward L-alanine 3-sulfinic acid at pH 8.0, and was also very active toward L-aspartic acid (Km = 1.6 mM). Ratios of activities for L-aspartic acid and L-cysteine were essentially constant during the purification of the enzyme. Evidence based on substrate specificity, enzyme inhibition, and physicochemical properties indicates that cytosolic cysteine aminotransferase is identical with cytosolic aspartate aminotransferase (L-aspartate: 2-oxoglutarate aminotransferase, EC 2.6.1.1).

Animals↗

PubMedDate not supplied

Interferon treatment in patients with chronic hepatitis C with normal alanine-aminotransferase activity.

BACKGROUND/AIMS: Chronic hepatitis C virus carriers may have repeatedly normal alanine aminotransferase activity despite detectable viremia and histological hepatitis. We aimed to evaluate the effect of interferon treatment in these cases. METHODOLOGY: Twelve patients with persistently normal alanine aminotransferase levels at least 6 months before therapy were treated with recombinant interferon (IFN)alpha-2b for 6 months, totaling 840 MU in amount. Alanine aminotransferase levels were measured monthly during treatment and after treatment withdrawal, and HCV-RNA levels were measured by polymerase chain reaction before treatment, and 6 and 12 months after treatment withdrawal. RESULTS: At treatment withdrawal, HCV-RNA levels had significantly decreased and HCV-RNA disappeared in 9 of the 12 patients by polymerase chain reaction. At 6 months after treatment withdrawal, HCV-RNA reappeared in 6 of the 9 patients whose HCV-RNA was negative at treatment withdrawal. Over all, only 4 of the 12 patients (33%) were sustained virological responders (HCV-RNA is negative more than 6 months after treatment withdrawal). Pre-treatment HCV-RNA levels in a sustained virological responder was significantly lower than that of transient and non-responders (4.9 +/- 1.6 vs. 7.7 +/- 1.6 log10[copies/ml], p < 0.05). Of 8 patients who did not achieved sustained virological response, alanine aminotransferase levels had transiently increased above normal during treatment in one patient and after treatment withdrawal in 6 patients; however, in the remaining one patient abnormal values have continued from 8 months after treatment withdrawal till now for 24 months. CONCLUSIONS: In patients with chronic hepatitis C with normal alanine aminotransferase levels, the response to interferon therapy was by no means satisfactory. However, if it would be used in cases with the lower pre-treatment HCV-RNA levels with careful attention to a transient alanine aminotransferase elevation, the more a sustained virological response might be expected.

Adult↗

PubMed2001

Evaluation of aminotransferase elevations in a bodybuilder using anabolic steroids: hepatitis or rhabdomyolysis?

The use of anabolic steroids among competitive athletes, particularly bodybuilders, is widespread. Numerous reports have noted "hepatic" dysfunction secondary to anabolic steroid use based on elevated serum aminotransferase levels. The authors' objective was to assess whether primary care physicians accurately distinguish between anabolic steroid-induced hepatotoxicity and serum aminotransferase elevations that are secondary to acute rhabdomyolysis resulting from intense resistance training. Surveys were sent to physicians listed as practicing family medicine or sports medicine in the yellow pages of seven metropolitan areas. Physicians were asked to provide a differential diagnosis for a 28-year-old, anabolic steroid-using male bodybuilder with an abnormal serum chemistry profile. The blood chemistries showed elevated aspartate aminotransferase (AST), alanine aminotransferase (ALT), and creatine kinase (CK) levels, and normal gamma-glutamyltransferase (GGT) levels. In the physician survey (n = 84 responses), 56% failed to mention muscle damage or muscle disease as a potential diagnosis, despite the markedly elevated CK level of the patient. Sixty-three percent indicated liver disease as their primary diagnosis despite normal GGT levels. Prior reports of anabolic steroid-induced hepatotoxicity that were based on aminotransferase elevations may have overstated the role of anabolic steroids. Correspondingly, the medical community may have been led to emphasize anabolic steroid-induced hepatotoxicity and disregard muscle damage when interpreting elevated aminotransferase levels. Therefore, when evaluating enzyme elevations in patients who use anabolic steroids, physicians should consider the CK and GGT levels as essential elements in distinguishing muscle damage from liver damage.

Adult↗

PubMed1976

Some properties of mitochondrial and cell sap alanine aminotransferase of the rat heart.

Alanine aminotransferase activity is present in mitochondria and the cell sap fraction of the rat myocardium. As distinct from the cell sap form, mitochondrial alanine aminotransferase was significantly inhibited by chloride ions, maleate and incubation medium temperatures of over 40 degrees C. Activity of the cell sap enzyme was inhibited by phosphate and stimulated by temperatures of over 40 degrees C. The pH optimum for cell sap alanine aminotransferase was in the region of 8, while for the mitochondrial enzyme it had a wider range (pH 7.3-8.2). D,L-penicillamine, and antagonist of vitamin B6, inhibited alanine aminotransferase activity equally in intact and tritonized mitochondria and in the cell sap fraction. The activity of mitochondrial and cell sap alanine aminotransferease rose in correlation to the stage of ontogenesis, the maximum increase being observed in the cell sap fraction 14-20 days after birth. The addition of coenzyme to the incubation medium did not affect the activity of either mitochondrial or cell sap alanine aminotransferase. The results indicate that there are two different alanine aminotransferase enzymes in the rat heart, with different intracellular localizations and probably with different regulative functions.

Alanine Transaminase↗

PubMed2002

Alanine aminotransferase deficiency in a hepatitis B surface antigen positive patient presenting with acute hepatitis.

This report presents a hepatitis B surface antigen positive case presenting with acute hepatitis and with findings of low serum alanine aminotransferase in contrast to very high levels of aspartate aminotransferase. A 64 year-old female patient was admitted to our hospital with fatigue and jaundice. Hepatitis B surface antigen was positive. During follow up, aspartate aminotransferase levels remained very high, while alanine aminotransferase levels continued to be extremely low. Additionally, all of the patients five daughters had low alanine aminotransferase levels. The clinical importance of alanine aminotransferase deficiency is still unclear.

Journal Article↗

PubMed1977

Interaction of glucocorticoid hormones and cyclic nucleotides in induction of tyrosine aminotransferase in cultured hepatoma cells.

Reproducible induction of the enzyme tyrosine aminotransferase by dibutyryl cAMP (Bt2cAMP) in a line of HTC hepatoma cells in suspension culture requires that the cells be preinduced with dexamethasone, a synthetic glucocorticoid which itself induces tyrosine aminotransferase. Concentrations of dexamethasone that do not induce tyrosine aminotransferase fail to support Bt2cAMP induction, removal of the steroid from the medium leads to a loss of the Bt2cAMP effect, and an HTC cell line whose aminotransferase is not steroid-inducible does not respond to the cyclic nucleotide. We show that the further induction of tyrosine aminotransferase by Bt2cAMP in dexamethasone-treated cells is due to an increased rate of enzyme synthesis. The cyclic nucleotide has no effect on aminotransferase synthesis in cells grown in the absence of steroid. Several lines of evidence suggest that dexamethasone acts at a step beyond the activation of protein kinase by cAMP: (a) basal levels of cAMP are not altered by growth of HTC cells in dexamethasone; (b) accumulation of cAMP from the medium is not enhanced; (c) the glucocorticoid does not induce cAMP-dependent protein kinase in HTC cells; and (d) there is no augmentation of cAMP binding to the regulatory protein, nor is there any change in cAMP activation of protein kinase caused by growth in dexamethasone. These results help define a system that should be useful in studying the interaction of cyclic nucleotides and steroid hormones.

Bucladesine↗

PubMed1977

Role of coenzyme in aminotransferase turnover.

The role of coenzyme in determining intracellular contnet of pyridoxal enzymes was assessed by analyzing effects of pyridoxine deficiency on the rapidly degraded, readily dissociable tyrosine aminotransferase (EC 2.6.1.5) and the slowly degraded, nondissociable alanine aminotransferase (EC 2.6.1.2) of rat liver. Synthesis of the tyrosine enzyme was reduced, leading to a decreased amount of this enzyme, much of which was present as active apoenzyme. Synthesis of alanine aminotransferase was unchanged but much of this enzyme was present as an inactive apoenzyme which retained immunological reactivity. Degradation rates of both enzymes (t1/2 about 1.5 h, tyrosine aminotransferase; about 3 days, alanine aminotransferase) were not changed in pyridoxine deficiency. Hence, interaction with coenzyme is not a significant determinant in intracellular degradation of these aminotransferases. Coenzymes dissociation and intracellular stability probably reflect structural features of the proteins which determine both properties.

Alanine Transaminase↗

PubMed1975

Effects of 6- and 8-substituted analogs of adenosine 3':5'-monophosphate on phosphoenolpyruvate carboxykinase and tyrosine aminotransferase in hepatoma cell cultures.

A variety of 6- and 8-substituted analogs of cAMP (cyclic adenosine 3:5-monophosphate) have been tested for their ability to increase activity of tyrosine aminotransferase (EC 2.6.1.5) in cultured Reuber H35 hepatoma cells. Some analogs, particularly the 8-thio-substituted ones, produced effects approximately equivalent to those generated by N-6, O2'-dibutyryl cAMP. In contrast, cAMP and its O-2-monobutyryl derivative were relatively ineffective even at very high concentrations, whereas three other analogs actually depressed the activity of the aminotransferase. Changes in enzyme activity generated by the various analogs were paralleled closely by changes in the relative rate of aminotransferase synthesis. An excellent correlation was found to exist between the ability of any given analog to influence the activity of tyrosine aminotransferase and that of phosphoenolpyruvate carboxykinase (EC 4.1.1.32). A similar correlation was found to exist between the ability of various analogs to evelate the activity of these enzymes and to inhibit reversibly the growth of H35 cells. Only one of five inhibitors of cAMP phosphodiesterase activity tested produce any increase in aminotransferase activity when added alone. All of the 6- and 8-substituted analogs tested, including noniducers, stimulated f1 histone phosphorylation in crude rat liver extracts with approximately equal potencies. On the other hand, dibutyryl cAMP was only a weak activator of protein kinase in vitro, even though it is a potent enzyme inducer. A possible resolution of this apparent discrepancy has been provided by preliminary analyses of site-specific f1 histone phosphorylation in whole cells. Only compounds active as aminotransferase inducers are capable of stimulating phosphorylation of the serine-37 residue of endogenous f1 histone (3- to 10-fold).

Animals↗

PubMed1986

Is cyclic AMP dependent protein kinase responsible for the in vivo phosphorylation of tyrosine aminotransferase?

Undegraded tyrosine aminotransferase was purified to near homogeneity from rat liver and was confirmed to be a substrate for the beef heart cyclic AMP dependent protein kinase catalytic subunit. Specific antibody was used to quantitate the amount of phosphate incorporated into the enzyme. Phosphate incorporation was maximal at a catalytic subunit to tyrosine aminotransferase molar ratio of 7:1 using 200 microM ATP for 30 to 60 min at 30 degrees C. Phospho-peptide maps of tyrosine aminotransferase phosphorylated in vitro by the catalytic subunit were compared with those of amino-transferase immunoprecipitated from 32P labeled cells treated with and without 8-Br cAMP. Whereas the phospho-peptide maps of tyrosine aminotransferase isolated from cells treated with and without 8-Br cAMP were identical, differences were observed in the peptide map of tyrosine aminotransferase phosphorylated in vitro and in vivo. These results were taken to indicate that the catalytic subunit is not responsible for tyrosine aminotransferase phosphorylation in vivo.

8-Bromo Cyclic Adenosine Monophosphate↗