scholarly journals Molecular analysis of the role of two aromatic aminotransferases and a broad-specificity aspartate aminotransferase in the aromatic amino acid metabolism ofPyrococcus furiosus

Archaea ◽  
2002 ◽  
Vol 1 (2) ◽  
pp. 133-141 ◽  
Author(s):  
Donald E. Ward ◽  
William M. de Vos ◽  
John van der Oost
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Aspartate Aminotransferase ◽  
Pyrococcus Furiosus ◽  
Aromatic Amino Acids ◽  
Significant Activity ◽  
Transamination Reaction ◽  
Genes Encoding ◽  
Branched Chain ◽  
Aromatic Amino Acid Metabolism

The genes encoding aromatic aminotransferase II (AroAT II) and aspartate aminotransferase (AspAT) fromPyrococcus furiosushave been identified, expressed inEscherichia coliand the recombinant proteins characterized. The AroAT II enzyme was specific for the transamination reaction of the aromatic amino acids, and uses α-ketoglutarate as the amino acceptor. Like the previously characterized AroAT I, AroAT II has highest efficiency for phenylalanine (kcat/Km= 923 s–1mM–1). Northern blot analyses revealed that AroAT I was mainly expressed when tryptone was the primary carbon and energy source. Although the expression was significantly lower, a similar trend was observed for AroAT II. These observations suggest that both AroATs are involved in amino acid degradation. Although AspAT exhibited highest activity with aspartate and α-ketoglutarate (kcat~105 s–1), it also showed significant activity with alanine, glutamate and the aromatic amino acids. With aspartate as the amino donor, AspAT catalyzed the amination of α-ketoglutarate, pyruvate and phenylpyruvate. No activity was detected with either branched-chain amino acids or α-keto acids. The AspAT gene (aspC) was expressed as a polycistronic message as part of thearooperon, with expression observed only when the aromatic amino acids were absent from the growth medium, indicating a role in the biosynthesis of the aromatic amino acids.

scholarly journals Aspartate: 2-oxoglutarate aminotransferase from trichomonas vaginalis. Identity of aspartate aminotransferase and aromatic amino acid aminotransferase

1985 ◽  
Vol 232 (3) ◽  
pp. 689-695 ◽  
Author(s):  
P N Lowe ◽  
A F Rowe
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Aspartate Aminotransferase ◽  
Aromatic Amino Acid ◽  
Trichomonas Vaginalis ◽  
Aromatic Amino Acids ◽  
Significant Activity ◽  
Irreversible Inhibitor ◽  
Competitive Substrate ◽  
Aromatic Amino Acid Aminotransferase

Aspartate: 2-oxoglutarate aminotransferase from the anaerobic protozoon Trichomonas vaginalis was purified to homogeneity and characterized. It is a dimeric protein of overall Mr approx. 100000. Only a single isoenzyme was found in T. vaginalis. The overall molecular and catalytic properties have features in common with both the vertebrate cytoplasmic and mitochondrial isoenzymes. The purified aspartate aminotransferase from T. vaginalis showed very high rates of activity with aromatic amino acids as donors and 2-oxoglutarate as acceptor. This broad-spectrum activity was restricted to aromatic amino acids and aromatic 2-oxo acids, and no significant activity was seen with other common amino acids, other than with the substrates and products of the aspartate: 2-oxoglutarate aminotransferase reaction. Co-purification and co-inhibition, by the irreversible inhibitor gostatin, of the aromatic amino acid aminotransferase and aspartate aminotransferase activities, in conjunction with competitive substrate experiments, strongly suggest that a single enzyme is responsible for both activities. Such high rates of aromatic amino acid aminotransferase activity have not been reported before in eukaryotic aspartate aminotransferase.

In Vitro adsorption Spectrum of Plasma Amino Acids by Coated Charcoal Hemoperfusion

1983 ◽  
Vol 6 (5) ◽  
pp. 267-270 ◽  
Author(s):  
Z.Q. Shi ◽  
T.M.S. Chang
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Hepatic Coma ◽  
Aromatic Amino Acids ◽  
Molar Ratio ◽  
Adsorption Experiment ◽  
Preferential Adsorption ◽  
Branched Chain ◽  
Charcoal Hemoperfusion

In order to clarify wether coated charcoal hemoperfusion is capable of normalizing amino acid disturbances in hepatic coma, in vitro adsorption and in vitro hemoperfusion studies were carried out. We have found that collodion-coated activated charcoal beads preferentially removed much more aromatic acids (AAA) than branched chain amino acids (BCAA). In the in vitro adsorption experiment with 50 μM amino acid standards aqueous solution, 99% of AAAs were removed by charcoal while only 50 to 81% of BCAAs were removed. As the concentration of amino acids in solution was doubled from μM to 100 μM, BCAA removal was halved while about 90% of AAA was still being removed. In vitro hemoperfusion with heparinized blood from hepatic failure rats, the clearance and the removal of AAAs were significantly greater than those of BCAAs. Consequently, the molar ratio of BCAA over AAA was markedly improved from the initial 1.09 to 3.87 after 60 min of hemoperfusion. Thus, we have demonstrated the preferential adsorption of aromatic amino acids by collodion-coated charcoal beads. The correction of BCAA/AAA molar ratio is also demonstrated.

Amino Acid Metabolism in Plants. III. Purification and Some Properties of a Multispecific Aminotransferase Isolated from Bushbean Seedlings (Phaseolus vulgaris L.)

1972 ◽  
Vol 50 (7) ◽  
pp. 813-829 ◽  
Author(s):  
J. C. Forest ◽  
F. Wightman
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Aspartic Acid ◽  
Aspartate Aminotransferase ◽  
Soluble Protein ◽  
Specific Activity ◽  
Aromatic Amino Acids ◽  
Total Activity ◽  
Aminotransferase Activity ◽  
Aromatic Aminotransferase

The development of aromatic aminotransferase activity was examined in cotyledons, roots, and shoots of bushbean seedlings growing under light or dark conditions for the first 2 weeks after germination. All three aromatic amino acid – α-ketoglutarate aminotransferase activities were found to have similar patterns of development in comparable organs grown under the two environmental conditions, and the changes in levels of activity appeared unrelated to variations in the endogenous amounts of free aromatic amino acids in the organs of these seedlings. The highest total activity for all three transamination reactions was found in the shoots of light-grown seedlings after 14 days, whereas the aminotransferases showing highest specific activity were found in roots of both kinds of seedlings after 8 days of growth. The intracellular distribution of the three aromatic aminotransferase activities and of aspartate aminotransferase activity was investigated by differential centrifugation of root homogenates. Only a total of 10% of these two activities was found in the two particulate fractions; the soluble protein in the final supernatant fraction accounted for almost 90% of the total aromatic and aspartate aminotransferase activities.The aromatic aminotransferase in the soluble protein fraction from seedling roots was purified about 600-fold by pH precipitation, ammonium sulfate fractionation, and Sephadex chromatography, and the recovery obtained was 30–35% based on total activity. It was observed that the specific activity for aspartate–α-ketoglutarate aminotransferase increased proportionally to the increase in aromatic aminotransferase activities during the different steps of purification. Gel electrophoresis of the purified fraction revealed only one protein band which corresponded to the product-specific stained band for the three aromatic aminotransferase activities assayed on other gels. The molecular weight of the purified aminotransferase was found to be about 128 000 daltons and its Stokes radius was calculated to be 43 ± 3 Å. The pH optima for the three aromatic aminotransferase activities and for aspartate aminotransferase activity were all found to be 8.5. The purified enzyme showed no specific requirement for pyridoxal phosphate and an examination of its amino acid substrate specificity revealed that it was able to catalyze transamination of L-aspartic acid, L-phenylalanine, L-tyrosine, and L-tryptophan when α-ketoglutarate was provided as amino group acceptor. The enzyme was also found to catalyze transamination of L-glutamic acid when oxaloacetate was used as amino group acceptor, but neither pyruvate nor glyoxylate were utilized as amino acceptors for transamination of any of the amino acids examined. The enzyme was found to catalyze transamination of aspartic acid with much greater velocity than its rate of reaction with any of the three aromatic amino acids, and the inclusion of aspartic acid in a reaction medium at equimolar concentration with any one of the three aromatic amino acids resulted in strong inhibition of the aromatic aminotransferase activity of the enzyme. All the evidence indicates that the soluble protein fraction purified from bushbean roots contained only one aminotransferase which was able to catalyze the transamination of five L-amino acids. The demonstration of the substrate multispeciftcity of this pure enzyme represents the first evidence for a multispecific aminotransferase in plants.

A Comparison of the Effects of Intravenous Infusion of Individual Branched-Chain Amino Acids on Blood Amino Acid Levels in Man

1981 ◽  
Vol 60 (1) ◽  
pp. 95-100 ◽  
Author(s):  
S. Eriksson ◽  
L. Hagenfeldt ◽  
J. Wahren
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Intravenous Infusion ◽  
Serum Insulin ◽  
Aromatic Amino Acids ◽  
Branched Chain Amino Acids ◽  
Blood Concentrations ◽  
Branched Chain Amino Acid ◽  
Amino Acid Levels ◽  
Branched Chain

1., Intravenous infusions of l-valine (600 μmol/min), l-isoleucine (150 μmol/min), l-leucine (300 μmol/min) and a mixture of the three branched-chain amino acids (70% l-leucine, 20% l-valine, 10% l-isoleucine; 270 μmol/min) were given to four groups of healthy volunteer subjects. Whole-blood concentrations of amino acids and glucose and serum insulin were measured before and during the infusions. 2. Valine and isoleucine infusions resulted in twelve- and six-fold increases in the respective amino acid. During valine infusion, tyrosine was the only amino acid for which a decrease in concentration was seen (25%, P < 0.05). With isoleucine administration, no significant changes were found. In contrast, leucine infusion (during which the leucine concentration rose about sixfold) was accompanied by significant decreases in tyrosine (35%), phenylalanine (35%), methionine (50%), valine (40%) and isoleucine (55%). The arterial glucose concentration fell slightly (5%) and the insulin concentration increased 20% during leucine infusion. 3. Infusion of the mixture of the three branched-chain amino acids resulted in marked decreases in tyrosine (50%), phenylalanine (50%) and methionine (35%). The decreased amino acid levels remained low for 2 h after the end of the infusion. 4. The present findings demonstrate that intravenous infusion of leucine (not infusion of valine or isoleucine) results in marked reductions in the concentrations of the aromatic amino acids and methionine. Infusion of a mixture of the three branched-chain amino acids gives results similar to those obtained with leucine infusion alone. Thus a mixed branched-chain amino acid solution with leucine as its main constituent seems to be the best alternative in the treatment of patients with hepatic cirrhosis and encephalopathy.

Genetic Characterization of the Major Lactococcal Aromatic Aminotransferase and Its Involvement in Conversion of Amino Acids to Aroma Compounds

1999 ◽  
Vol 65 (11) ◽  
pp. 4873-4880 ◽  
Author(s):  
Liesbeth Rijnen ◽  
Sophie Bonneau ◽  
Mireille Yvon
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Physiological Role ◽  
Aromatic Amino Acids ◽  
Aroma Compounds ◽  
Gene Inactivation ◽  
Sequence Homologies ◽  
Branched Chain ◽  
Aromatic Aminotransferase

ABSTRACT In lactococci, transamination is the first step of the enzymatic conversion of aromatic and branched-chain amino acids to aroma compounds. In previous work we purified and biochemically characterized the major aromatic aminotransferase (AraT) of a Lactococcus lactis subsp. cremoris strain. Here we characterized the corresponding gene and evaluated the role of AraT in the biosynthesis of amino acids and in the conversion of amino acids to aroma compounds. Amino acid sequence homologies with other aminotransferases showed that the enzyme belongs to a new subclass of the aminotransferase I subfamily γ; AraT is the best-characterized representative of this new aromatic-amino-acid-specific subclass. We demonstrated that AraT plays a major role in the conversion of aromatic amino acids to aroma compounds, since gene inactivation almost completely prevented the degradation of these amino acids. It is also highly involved in methionine and leucine conversion. AraT also has a major physiological role in the biosynthesis of phenylalanine and tyrosine, since gene inactivation weakly slowed down growth on medium without phenylalanine and highly affected growth on every medium without tyrosine. However, another biosynthesis aromatic aminotransferase is induced in the absence of phenylalanine in the culture medium.

Effect of Hemodiabsorption and Sorbent-Based Pheresis on Amino Acid Levels in Hepatic Failure

2000 ◽  
Vol 23 (6) ◽  
pp. 375-388 ◽  
Author(s):  
J. Steczko ◽  
K.C. Bax ◽  
S.R. Ash
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Hepatic Failure ◽  
High Performance ◽  
Aromatic Amino Acids ◽  
Dialysis Unit ◽  
Amino Acid Profiles ◽  
Amino Acid Levels ◽  
Pre Treatment ◽  
Branched Chain

Changes in plasma amino acid concentrations were measured in patients with hepatic failure during extracorporeal hemodiabsorption (using the Liver Dialysis Unit, “the Unit”) or hemodiabsorption plus sorbent-based pheresis treatment (using the Liver Dialysis Plasmafilter Unit, “the PF-Unit”) Systems. Eight patients with hepatic failure, grade 3 or 4 encephalopathy, elevated bilirubin and/or creatinine levels and respiratory or renal failure were treated for 1–3 days with the Unit alone. Three of these were also treated with the Unit containing 10 g of BCAA in the sorbent suspension. Four patients with hepatic failure treated with the PF Unit also had 10 g of branched chain amino acid (BCAA) added to the sorbents of the Unit portion of this device. Pre- and post-plasma samples were drawn and high performance liquid chromatography (HPLC) was used to separate and detect amino acids in the plasma. Both the Unit and the PF-Unit have the capability to selectively remove various amino acids, especially aromatic amino acids (AAA). The pre-treatment amino acid profiles of plasma were typical for hepatic failure, with abnormally high levels of phenylalanine, tyrosine, tryptophan, and methionine and decreased levels of valine, leucine and isolucine. The average pre-treatment Fischer ratio (BCAA/AAA) for both Unit and PF-Unit patients was 1.43 (±0.58). Treatments by both systems resulted in an increase of BCAA levels in blood and concomitant decrease of AAA levels, with an average Fischer ratio improvement of 30–38% for the Unit and PF-Unit without BCAA. The Fischer ratio improved by 90% (average) for the Unit with BCAA. Levels of many other amino acids (such as alanine, glycine, proline or lysine) increased during both Unit and PF-Unit treatments. The removal of strongly protein-bound toxin and amino acids such as tryptophan and sulphydryl amino acids was more effective by the PF-Unit. Both the Unit and the PF-Unit have the unique capability to remove toxic aromatic amino acids while increasing BCAA levels in patient. The increase in many amino acid levels may be related to the removal of toxins that interfere with normal amino acid metabolism. The addition of the PF module improves the removal of bilirubin and similarly protein-bound chemicals. Changes in amino acid profiles by the Unit and the PF-Unit contrast markedly with other extracorporeal devices.

scholarly journals A Hydrolase from Lactobacillus sakei Moonlights as a Transaminase

2013 ◽  
Vol 79 (7) ◽  
pp. 2284-2293 ◽  
Author(s):  
Quirin Sinz ◽  
Simone Freiding ◽  
Rudi F. Vogel ◽  
Wilfried Schwab
Keyword(s):  
Amino Acids ◽  
Amino Acid ◽  
Aromatic Amino Acids ◽  
Initial Step ◽  
Starter Cultures ◽  
Fermented Foods ◽  
Lactobacillus Sakei ◽  
Bacterial Strains ◽  
Content Type ◽  
Branched Chain

ABSTRACTEnzymatic transamination of amino acids yields α-keto acids and is the initial step for the production of volatile compounds that contribute to the sensory perception of fermented foods such as salami.Lactobacillus sakeiis one of the lactic acid bacterial strains commonly used in starter cultures. Although the genome sequence ofL. sakei23K lacks genes encoding typical branched-chain amino acid transaminases, transamination activity and the formation of amino acid-derived volatile metabolites could be demonstrated. A protein purified fromL. sakeiis held responsible for the transamination activity. By heterologous expression of the corresponding gene inEscherichia coli, we were able to characterize the transamination side activity of an enzyme annotated as a putative acylphosphatase (AcP). A transamination side activity of hen egg white lysozyme (HEWL) was also discovered. Both enzymes showed substrate specificity toward branched-chain and aromatic amino acids. AcP also accepted l-methionine. Activity was optimal at neutral pH for both enzymes, whereas AcP showed a significantly higher temperature optimum (55°C) than that of HEWL (37°C). Kinetic parameters revealed high affinity towardl-leucine for AcP (Km= 1.85 mM) and towardl-isoleucine for HEWL (Km= 3.79 mM). AcP seems to play a major role in the metabolism of amino acids inL. sakei.

Effects of Sorbent Suspension Dialysis on Plasma Amino Acid Levels in Cirrhotic Patients with Refractory Hepatic Encephalopathy

2002 ◽  
Vol 25 (10) ◽  
pp. 923-928 ◽  
Author(s):  
E. Bauer ◽  
A. Gendo ◽  
C. Madl ◽  
F. Garo ◽  
E. Roth ◽  
...  
Keyword(s):  
Amino Acids ◽  
Hepatic Encephalopathy ◽  
Amino Acid ◽  
Medical Treatment ◽  
Aromatic Amino Acids ◽  
Plasma Amino Acid ◽  
Clinical Grade ◽  
Cirrhotic Patients ◽  
Amino Acid Levels ◽  
Branched Chain

In cirrhotic patients, plasma amino acid levels are severely deranged. A decreased ratio of branched-chain to aromatic amino acids (Fischer ratio) has been implicated in the pathogenesis of hepatic encephalopathy. In this prospective study, we investigated the effects of extracorporeal detoxification on amino acid levels using a sorbent suspension dialysis system. Twenty patients with documented cirrhosis and hepatic encephalopathy grade II-III not responding to standard treatment were randomized to receive either six hours of sorbent dialysis and standardized conventional medical treatment or ongoing medical treatment alone. In contrast to previous uncontrolled studies, no significant effect on amino acid levels, Fischer ratio or clinical grade of hepatic encephalopathy was detected in either treatment group. In conclusion, a 6-hour treatment with sorbent dialysis did not significantly influence plasma levels of amino acids and did not ameliorate the clinical grade of hepatic encephalopathy.

scholarly journals Long-Term Carbohydrate-Containing Late-Evening Snack Significantly Improves the Ratio of Branched Chain Amino Acids to Aromatic Amino Acids in Adults with Liver Cirrhosis due to Hepatitis B

2021 ◽  
Vol 2021 ◽  
pp. 1-11
Author(s):  
Wei Hou ◽  
Zheng Lv ◽  
Jing Yang ◽  
Jing Wu ◽  
Zhong-ying Wang ◽  
...  
Keyword(s):  
Amino Acids ◽  
Liver Cirrhosis ◽  
Chronic Hepatitis ◽  
Amino Acid ◽  
Aromatic Amino Acids ◽  
Branched Chain Amino Acids ◽  
Plasma Amino Acid ◽  
Amino Acid Profiles ◽  
Late Evening ◽  
Branched Chain

Background. The liver is the primary organ for amino acid metabolism, and metabolic disorder of amino acids is common in liver disease. However, the characteristics of plasma amino acid profiles in patients with HBV-related cirrhosis and the impacts of late-evening snack (LES) on cirrhosis are unclear. Objectives. To investigate the characteristics of plasma amino acid profiles in patients with HBV-related chronic hepatitis, cirrhosis, and the effects of late-evening snacks on plasma amino acid profiles. Methods. 86 patients with HBV-related cirrhosis and eighty patients with chronic hepatitis B were included in this study. The plasma amino acid profiles were measured by the amino acid analyzer. Patients were randomly divided into two groups, of which the liver cirrhosis group was to receive daily LES ( n = 43 ) or non-LES ( n = 43 ) for 6 months. Plasma amino acid profiles and biochemical parameters were measured in both groups at baseline and after 1, 3, and 6 months. Results. Compared to healthy controls, the plasma concentration in the liver cirrhosis group of threonine, serine, glycine, glutamine, cysteine, tyrosine, phenylalanine, arginine, and methionine increased significantly ( P < 0.05 ), while the ratio of branched chain amino acids (BCAA) to aromatic amino acids (AAA) decreased significantly ( P < 0.05 ). A carbohydrate-predominant LES treatment resulted in a significant increase in BCAA/AAA and decrease in the level of ammonia and glutamine compared with baseline after 6 months of supplementation ( P < 0.05 ). Patients with Child-Pugh B and C are more responsive to changes in amino acid profiles than those with Child-Pugh A. Conclusions. The application of an LES carbohydrate module for six months in liver cirrhosis patients was associated with increased BCAA/AAA and decreased level of ammonia. Patients with Child-Pugh B and C grades were the most beneficial population.

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