02 Amino Acid Deamination

Stage I Converts an Amino Acid to a Keto Acid

(1) Transimination: The amino acid's nucleophilic amino group attacks the enzyme-PLP Schiff base carbon atom in a transimination reaction to form an amino acid-PLP Schiff base (aldimine), with release of the enzyme's Lys amino group. [This Lys is then free to act as a general base at the active site] (2) Tautomerization: The amino acid-PLP Schiff base tautomerizes to an α-keto acid-PMP Schiff base (ketimine) by the 1. active site Lys-catalyzed removal of the amino acid α-hydrogen and 2. protonation of PLP atom C4' via a resonance-stabilized carbanion intermediate. (3) Hydrolysis: The α-keto acid-PMP Schiff base is hydrolyzed to PMP and an α-keto acid.

PLP

-Enzymes that catalyze transamination, called aminotransferases or transaminases, require the coenzyme pyridoxal-5'-phosphate (PLP) -PLP is a derivative of pyridoxine (vitamin B6) -The coenzyme is covalently attached to the enzyme via a Schiff base (imine) linkage

Amino Acid Deamination

-Free amino acids originate from: 1. the degradation of cellular proteins and 2. the digestion of dietary proteins -substances are absorbed by the intestinal mucosa and transported through the bloodstream to be absorbed by other tissues -further degradation of amino acids takes place intracellularly, the α-amino group is removed. -the amino group is converted to ammonia, incorporated into urea for excretion -The remaining carbon skeleton (α-keto acid) of the amino acid can be broken down to other compounds

GDH: Glutamate Can Be Oxidatively Deaminated

-Glutamate can be oxidatively deaminated by glutamate dehydrogenase (GDH)→ammonia and regenerating α-ketoglutarate -Glutamate dehydrogenase, a mitochondrial enzyme,can accept either NAD+ or NADP+ as its redox coenzyme. -Oxidation occurs with transfer of a hydride ion from glutamate's Cα to NAD(P)+, forming α-iminoglutarate, which is hydrolyzed to α-ketoglutarate and ammonium ion:

Transaminases Use PLP to Transfer Amino Groups

-Most amino acids are deaminated by transamination, the transfer of the amino group to an α-keto acid to yield the α-keto acid of the original amino acid and a *new amino acid. -α-ketoglutarate is the predominant amino group acceptor, producing glutamate and the new α-keto acid -Glutamate's amino group can be transferred to oxaloacetate in a second transamination reaction, yielding aspartate and reforming α-ketoglutarate -The presence of transaminases in muscle and liver cells makes them useful markers of tissue damage. -The concentrations of these enzymes in the blood increase after a heart attack, when damaged heart muscle leaks its intracellular contents. -Liver damage is also monitored by SGOT and SGPT levels -Assays of the enzymes' activities in the blood are the basis of the commonly used clinical measurements known as SGOT (serum glutamate-oxaloacetate transaminase, also known as aspartate transaminase, AST) and SGPT (serum glutamate-pyruvate transaminase, or alanine transaminase, ALT).

GDH regulation

-The enzyme is allosterically inhibited by GTP and NADH (signaling abundant metabolic energy) and activated by ADP and NAD+ (signaling the need to generate ATP) -α-ketoglutarate is an intermediate of the citric acid cycle→activation of glutamate dehydrogenase can stimulate flux through the citric acid cycle,→leading to increased ATP production by oxidative phosphorylation. -The equilibrium position of the glutamate dehydrogenase reaction (ΔG°'≈ 30 kJ · mol-1, standard conditions) favors glutamate synthesis, the reverse of the reaction shown. -The ammonia liberated in the GDH reaction as shown previously is eventually excreted in the form of urea. (Under physiological conditions, the enzyme was thought to function close to equilibrium so changes in ammonia concentration could cause a shift in equilibrium toward glutamate synthesis to remove the excess ammonia.)

Stage II Converts an α-Keto Acid to an Amino Acid

To complete the aminotransferase's catalytic cycle, the coenzyme must be converted from PMP back to the enzyme-PLP Schiff base. (3') PMP reacts with an α-keto acid to form a Schiff base. (2') The α-keto acid-PMP Schiff base tautomerizes to form an amino acid-PLP Schiff base. (1') The ε-amino group of the active site Lys residue attacks the amino acid-PLP Schiff base in a transimination reaction to regenerate the active enzyme-PLP Schiff base and release the newly formed amino acid.