Branched-Chain Amino Acid Metabolism

Harper, Alfred E.; Miller, Robert H.; Block, Kevin P.

doi:10.1146/annurev.nu.04.070184.002205

Cited by 983 publications

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“…The activity state of mammalian branched-chain ␣-ketoacid dehydrogenase (BCKDH) 1 is highly regulated according to the physiological requirements for either degradation or conservation of branched-chain amino acids derived from cellular and dietary protein (1,2). This regulation is primarily exerted through reversible phosphorylation of the E1␣ subunit by BCKDH kinase.…”

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confidence: 99%

Roles of Amino Acid Residues Surrounding Phosphorylation Site 1 of Branched-chain α-Ketoacid Dehydrogenase (BCKDH) in Catalysis and Phosphorylation Site Recognition by BCKDH Kinase

Hawes

Schnepf

Jenkins

et al. 1995

Journal of Biological Chemistry

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Branched-chain ␣-ketoacid dehydrogenase is regulated by reversible phosphorylation of serine 293 (site 1) on the E1␣ subunit. Alanine-scanning mutagenesis was used to examine the roles of residues surrounding serine 293 in catalysis by the dehydrogenase and in substrate recognition by branched-chain ␣-ketoacid dehydrogenase kinase. Alanine substitution of serine 293 resulted in a 10-fold increased K m for ␣-ketoisovalerate, a less increased (2.8-fold) K m for ␣-ketoisocaproate, but no change in V max or the K m for thiamine pyrophosphate. Alanine substitutions of arginine 288, histidine 292, and aspartate 296, residues highly conserved among ␣-ketoacid dehydrogenases, resulted in inactive enzymes. Each of the inactive E1 mutants bound to the E2 core subunit with equal affinity as wild-type E1, and each produced circular dichroism spectra identical to that of wild-type E1. Two mutations, H292A and S293E, abolished the ability of E1 apoenzyme to reconstitute with thiamine pyrophosphate. Each alanine-substituted E1 was phosphorylated at site 1 by branched-chain ␣-ketoacid dehydrogenase kinase with similar rates, with the exception of the R288A mutant, which displayed no detectable phosphorylation. Thiamine pyrophosphate inhibited the phosphorylation of all mutant enzymes with the exception of H292A, the mutant E1 that did not bind thiamine pyrophosphate.The activity state of mammalian branched-chain ␣-ketoacid dehydrogenase (BCKDH) 1 is highly regulated according to the physiological requirements for either degradation or conservation of branched-chain amino acids derived from cellular and dietary protein (1, 2). This regulation is primarily exerted through reversible phosphorylation of the E1␣ subunit by BCKDH kinase. BCKDH kinase phosphorylates two serine residues on the E1␣ subunit, although inactivation of the dehydrogenase results exclusively from the phosphorylation of site 1, serine 293 in rat E1␣ (3-5). Furthermore, phosphorylation of site 2 occurs at a significantly slower rate (3-5), suggesting that the site 2 serine, serine 303 in rat E1␣, is a poor substrate either due to the primary or secondary structure surrounding this residue or the positioning of this serine residue relative to the kinase active site. BCKDH kinase is regulated in part through inhibition by branched-chain ␣-ketoacids and thiamine pyrophosphate, all of which are substrates for the BCKDH-catalyzed reaction (6, 7). Inhibition of the kinase by branched-chain ␣-ketoacids is believed to occur in vivo, resulting in a fully active BCKDH in the liver of rats fed a high protein diet (2). However, the exact molecular basis for these inhibitory effects is not clearly understood. BCKDH kinase has also been reported to be activated by its interaction with the E2 subunit of the BCKDH complex, although the molecular basis for this mode of regulation is also not understood (8, 9). Another important feature of BCKDH kinase is its high degree of substrate specificity, resulting in phosphorylation only of BCKDH E1␣ but not the highly homologous E1␣ su...

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confidence: 99%

Roles of Amino Acid Residues Surrounding Phosphorylation Site 1 of Branched-chain α-Ketoacid Dehydrogenase (BCKDH) in Catalysis and Phosphorylation Site Recognition by BCKDH Kinase

Hawes

Schnepf

Jenkins

et al. 1995

Journal of Biological Chemistry

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“…Consequently, levels of leucine were in excess of their minimum requirements (Table 1). Many amino acids, when fed in excess to broiler growing chickens cause symptoms of toxicity such as decreased feed intake, weight gain and increased mortality [8,15,29] . L-leucine in excess may cause toxicity in chicks fed low protein diets as reported by Farran et al [14,28] .…”

Section: Discussionmentioning

confidence: 99%

Effect of L-Leucine Supplementation on Growth Performance and Carcass Characteristics of Grower-Broiler Chickens Fed Low Protein Diets

Erwan¹,

Alimon²,

Sazili³

et al. 2009

American J. of Animal and Veterinary Sciences

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Problem statement: Supplementation of broiler diets with cristalline amino acids (i.e. lysine, ethionine and threonine) may support equal broiler growth and improve overall amino acids balance and enable a reduction in CP level of diets. Approach: A trial was conducted to evaluate the effect of supplemental L-leucine in diets containing recommended levels and low crude protein (20 and 18%, respectively) with constant metabolizable energy (3200 kcal kg −1 ) for broilers from 21-42 day of age. Six experimental diets were formulated with three levels of supplemental L-leucine, 0, 0.5 and 0.67% and two levels of crude protein. A total of 180 1 day-old Cobb broiler chickens were randomly divided into 36 experimental pens, 5 chickens in each pen, with each diet replicated 6 times. The dietary treatments were offered from 21-42 days of age. Feed intake, body weight gain and Feed Conversion Ratio (FCR) were measured on a weekly basis. At the end of the feeding trial the birds were slaughtered and carcass analyses conducted. Results: Feed intake, weight gain and FCR were not affected by increasing levels of L-leucine supplementation. Weight gain was significantly reduced (p<0.05), whereas feed intake and FCR were not significantly affected with decreasing dietary crude protein. A positive response in breast meat yield was achieved by the addition of L-leucine to levels up to 0.5% in the diet but a significant decrease was noted when the level reached 0.67% in diet. Supplementation of L-leucine significantly (p<0.05) decreased the relative weights of the liver and gizzard. However, the addition of L-leucine significantly reduced carcass weights when L-leucine was added at 0.67%. Lowering the dietary protein level also significantly reduced breast yield and carcass weight (p<0.05). However, abdominal fat, gizzard, liver and heart were not affected by protein level. Conclusion/Recommendations: It can be concluded that supplementation of L-leucine at levels up to 0.67% of the diet did not affect performance but deleteratious the carcass weight.

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“…With substrates A -L-leucine-and B-2-oxo glutarate-(and the corresponding apparent Michaelis constants KW {A) and KM(B)) in the almost complete ab sence of products and taking into account possible in hibitory effects of substrates (inhibitory constants and -Ki(B)), the velocity of the forward reaction (u) is given by formula [1]: Numerical fitting of the equation to the measured kinetic data was accomplished by means of a nonlinear regression procedure (NUN) from the SAS system (SAS Institute, Cary, NC) for personal computers using Marquardt's method, The provisional estimates for V'max» and needed for computation were de termined graphically according to Eq. Other results are means of two (deviation from mean < or ^10%) or means ± SD (number of determinations in parentheses).…”

Section: Enzyme Activity Testsmentioning

confidence: 99%

“…It is catalyzed by branched-chain L-amino acid aminotransferase (EC 2.6.1.42) (1,2). The enzyme has been purified to appar ent homogeneity from a number of mammalian tissues, e.g., from pig heart (3-5) and brain (6) and rat heart (7), liver (8), pancreas (9), and brain (10).…”

mentioning

confidence: 99%

Enzymatic Method for Determination of Branched-Chain Amino Acid Aminotransferase Activity

Schadewaldt

Hummel

Wendel

et al. 1995

Analytical Biochemistry

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A spectrophotom etric assay for th e determination of branched-chain L-amino acid am inotransferase activ ity is described. It is based on th e transamination of L-leucine in the presence of 2-oxoglutarate yielding 4-ethyl-2-oxopentanoate. The rate of formation of the branched-chain 2-oxo acid is specifically monitored in a coupled enzymatic reaction using NADf -dependent D*2-hydroxyisocaproate dehydrogenase from L actoba c illu s casei sap. p se u d o p la n ta ru m as coupling enzyme by m easuring the decrease in NADH absorbance at 334 nm. Optimized assay conditions are provided as evalu ated for the (iso)enzyme from rat heart, c 1995 Academic Press, Inc.

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Branched-Chain Amino Acid Metabolism

Cited by 983 publications

References 0 publications

Roles of Amino Acid Residues Surrounding Phosphorylation Site 1 of Branched-chain α-Ketoacid Dehydrogenase (BCKDH) in Catalysis and Phosphorylation Site Recognition by BCKDH Kinase

Roles of Amino Acid Residues Surrounding Phosphorylation Site 1 of Branched-chain α-Ketoacid Dehydrogenase (BCKDH) in Catalysis and Phosphorylation Site Recognition by BCKDH Kinase

Effect of L-Leucine Supplementation on Growth Performance and Carcass Characteristics of Grower-Broiler Chickens Fed Low Protein Diets

Enzymatic Method for Determination of Branched-Chain Amino Acid Aminotransferase Activity

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