경희대학교 의과대학·의학전문대학원

AMINO ACIDS METABOLISM

2019 년 1 학기

Text : Harper’s Biochemistry Refs: Devlin’s, Lippincott’s

Byung Kwan Jin, Ph.D.

Professor

Dept. of Biochemistry & Molecular Biology

School of Medicine Kyung Hee University

Ch. 27.

Biosynthesis

of the Nutritionally Nonessential Amino Acids

Ch. 28. Catabolism of Proteins & of Amino Acid

Nitrogen

Ch. 29. Catabolism of the

Carbon Skeletons

of Amino Acids

Ch. 30.

Conversion

of Amino Acids to Specialized Products

AMINO ACIDS METABOLISM

Car-bons

AMINO ACIDS METABOLISM

Car-bons

NH

Urea

Chapter 27.

Biosynthesis of the Nutritionally Nonessential

Amino Acids

Biomedical Importance

* Medical implications related to the Amino acid (AA) deficiency:

If nutritionally essential AAs are absent from the diet,

or are present in inadequate amounts

* Amino acid deficiency:

kwashiorkor

marasmus

scurvy

Nutritionally essential & Nutritionally nonessential AAs

* All 20 AAs are essential to ensure health.

•Nutritionally essential AAs: (8) must be present in the human diet.

•Nutritionally nonessential AAs: (12) need not be present in the diet

(Table 27-1)

• Nutritionally nonessential AAs have small number of enzymes required to

synthesize them

• Nutritionally nonessential AAs have short biosynthetic pathways

* Reactions and intermediates involved in the Biosynthesis (by human tissues) of

the 12 nutritionally nonessential 12 AAs

* Selected nutritional and metabolic disorders associated with metabolism

Chapter 27.

Biosynthesis of the Nutritionally Nonessential

Amino Acids

Carbons

Glutamate dehydrogenase

Glutamine synthetase

Aminotransferase

Carbons

NH

AA biosynthesis :

Figure 27-1 The glutamate dehydrogenase

reaction

Glutamate

- Reductive amidation of α-ketoglutarate by glutamate dehydrogenase

- 1

_{step in the biosynthesis of the ‘glutamate family’}

Figure 27-2 The

re-action

Glutamine

Alanine

:

Transamination

of pyruvate

Aspartate

:

Transamination

of oxaloacetate

Figure 27-4 The asparagine synthetase reaction

Asparagine

: Conversion of Aspartate to Asparagine

Amin o

donor

Figure 27-2 The glutamine synthetase reaction

Amin o

Figure 27-5 Serine biosynthesis (-AA, -amino acids; -KA, -keto

acids)

Serine

Oxidation of α-hydroxyl group of 3-phosphoglycerate to oxo acid   transamination  dephosphorylation  Ser

Figure 27-6 Formation of glycine from choline

Glycine

Glycine aminotransferase : Gly from glyoxylate and Glu or Ala

② From choline  betaine  Gly (Fig 27-6)

+ Glu or Ala

; Amino donor

Glycine aminotransferase

2

1

2

1

Glycine

① Glycine aminotransferase : Gly from glyoxylate and Glu or Ala ② From choline  betaine  Gly (Fig 27-6)

③ From Ser by serine hydroxymethyltransferase (Fig 27-7)

Figure 27-7

The serine hydroxymethyltransferase reac-tion

3

Figure 27-8 Biosynthesis of proline from glutamate

Proline

Proline

Cysteine

Cys (nutritionally nonessential AA) from Met (nutritionally essential AA) : Met  SAM  homocys-teine (Fig. 29-19)

L-Homecysteine + Ser  Cystathionine  L-Cys and homoserine (Fig. 27-9)

Figure 27-9 Conversion of homocysteine and serine to homoserine and cys-teine

SAM

Figure 27-10 The phenylalanine hydroxylase re-action

Tyrosine

① Phenylalanine hydroxylase : Phe to Tyr (Fig. 27-10)

② Catalysis by mixed function oxygenase : O₂ into Phe and water ③ Reducing power by tetrehydrobiopterin from NADPH

Figure 27-11 The prolyl (lysyl) hydroxylase re-action

Hydroxyproline and hydroxylysine

② no tRNA for hydroxylated AA ㅡ > neither dietary hydroxyproline nor hydroxylysine is

incorporated during protein synthesis ㅡ > Post-translationally, hydroxylation by prolyl hydroxylase and lysyl hydroxylase of tissues (skin, skeletal muscle…) (Fig. 27-11)

③ Hydroxylases (mixed-function oxygenases)

substrate (proline <lysine> -rich peptide, O₂, ascorbate, Fe2+ _{, and α-ketoglutarate}  succinate+ hydroxy-pro or hydroxy-lys

**

scurvy :

a proline (lysine)-rich peptide

lysine

Valine, Leucine, and Isoleucine:

nutritionally

①

② Thus, these α-keto acids can replace their AAs in the diet

Amino donor

Figure 27-12 Selenocysteine (top) and the reaction cat-alyzed

by selenophosphate systhetase (bottom)

Selenocysteine, the 21

_{Amino acids}

• SelenoCys in

active sites

of several enzymes: thioredoxin reductase, gluthathione

peroxi-dase..

• Tumorigenesis, atherosclerosis, selenium deficiency cardiomyopathy (Keshan disease)

Cys-teine

Chapter 28.

Catabolism of Proteins & of Amino Acid Nitrogen

* Biomedical Importance

①

How nitrogen of AAs is converted to

urea

and rare disorders that

accom-pany defects in urea biosynthesis

② Ammonia is highly toxic, tissues

convert ammonia to amide nitrogen

of

Gln (non-toxic)

 Subsquent deamination of Gln in

liver releases ammonia

 urea

(non-toxic)

③ Liver dysfunction (

간경화 , 간

염 ) elevated blood ammonia

levels  clinical signs and

symp-toms

NH

•Protein

turnover (degradation & synthesis)

occurs in all forms of life

① Humans turn over 1-2% of total body protein per day, principally muscle protein

② Liberated amino acids ~ 75%  reutilized

③ Excess amino acids  not stored but rapidly degraded nitrogen  urea

*Protease & peptidases degrade proteins to AAs

① Half-life t1/2 (susceptibility) of protein:

30 min to >150 hrs (half-lives of liver protein) ② PEST sequences: rapid degradation

- regions rich in proline(p), glutamate(E), serine(S), thronine(T)

③ Peptide hydrolysis: intracellular proteases 

Figure 28-1 Reactions involved in the attachment of ubiquitin (Ub) to pro-teins

⑤ Short half-lives, Abnormal and

Misfolded proteins : degraded in the cytosol

 requires ATP and

ubiquitin (present in

all eukaryotic cells)

 Ubs are attached by non-α-peptide bond

formed between carboxyl terminal of Ub and

α-amino groups of lysyl residues in target

protein.

 E1 Ub activating Enz.

E2 Ub transferase (ligase)

E3 Ub ligase (transferase)

•Amino terminal affect ubiquitination:

amino terminal Met or Ser

retards,

whereas

Asp or Arg

accelerates

ubiquitination

* Degradation of ubiquitin-tagged proteins

takes place in

proteasome

④ Extracellular, membrane-associated, long-lived proteins  degraded in lysosomes by ATP-inde-pendent processes Defects of Ubiquitina-tion; Angelman syndrome. Defect in E3 ligase; Hippel-Lindau syndrome

4

5

5

The multiple ubiquitin-tagging hypothesis (1981)

: Novel prize for the discovery of ubiquitin-mediated protein degradation (2004)

Chemistry – Technion (Israel)

Avram Hershko

Aaron Ciechanover1

UC, Irvine,

Figure 28-2 Interorgan amino acid exchange in normal post absorptive humans

*

Interorgan exchange maintains circulating levels of AAs

 Liver: the site of urea cycle enzymes necessary for disposal of excess nitrogen

* Ala & Gln : released from muscle into circulation

* Ala : vehicle of nitrogen transport in the plasma

extracted primarily by the liver

Figure 28-3 The glucose-alanine cycle Figure 28-4 Summary of amino acid exchange between organs immediately after feeding

•Interorgan exchange maintains circulating levels of AAs

 Ala: key gluconeogenic AA

 Rate of hepatic gluconeogenesis from Ala >>>>> all other AAs

 Branched-chain AAs (Leucine, Valine, Isoleucine) serve a special role in nitrogen metabolism, both in fasting state, when they provide brain with an energy source,

and after feeding, when they are extracted predominantly by muscles, having been spared by liver.

fasting state

•Animals convert α–amino nitrogen to varied end products

① Different animals excrete excess nitrogen as ammonia, uric acid, or urea

Fish : ammonotelic (excrete ammonia; highly toxic) Birds : uricotelic (excrete uric acid)

Humans : ureotelic (excrete nontoxic, water-soluble urea)

Biosynthesis of Urea

Four stages:

(1) Transamination,

(2) Oxidative deamination of Glu

(3) Ammonia transport

(4) Urea cycle

Figure 28-5 Overall flow of nitrogen

in amino acid catabolism

PLP in linkage to

aminotransferase

Biosynthesis of Urea

* Transamination transfers α-amino nitrogen to α-ketoglutarate, forming glutamate

① Transamination: α-amino acids and α-keto acids by aminotransferase (Fig. 28-6; Fig. 7-4; “ping-pong mechanism), except Lys, Thr, Pro, Hyp

② Coenzyme pyridoxal phosphate (PLP; A derivative of Vitamin B6, Fig. 44-12)

present at catalytic site of all aminotransferase

 enzyme-bound PLP serve as a carrier of amino groups

** remaining carbon “skeleton” of an AA is degraded by pathways (Chap. 29)

Figure 28-6 Transamination

1

1

2

Figure 28-7 Alanine aminotransferase and

glutamate aminotransferase

Biosynthesis of Urea

•Transamination transfers α-amino nitrogen to α-ketoglutarate, forming glutamate

• _③ Alanine aminotransferase : (Alanine-pyuvate aminotransferase)

glutamate aminotransferase : (Glutamate-α-ketoglutarate aminotransferase) (Fig. 28-7)

glutamate aminotransferase

Alanine aminotransferase

3

Biosynthesis of Urea

•Transamination transfers α-amino nitrogen to α-ketoglutarate, forming glutamate

• L-Glu is the only AA that undergoes oxidative deamination by glutamate dehydrogenase (GDH) (Fig. 28-8; Fig. 27-1)

The formation of ammonia from α-amino groups thus occurs mainly via the α-amino nitrogen of L-Glu

⑤ Serum levels of aminotransferases are elevated in some disease (Table 7-2): ALT (SGPT), AST (SGOT)

Figure 28-8

The L-glutamate dehydrogenase reaction

(Figure 27-1)

4

Biosynthesis of Urea

Four stages:

(1) Transamination,

(2) Oxidative deamination of Glu,

(3) Ammonia transport

(4) Urea cycle

Figure 28-5 Overall flow of nitrogen

in amino acid catabolism

Figure 28-8 The L-glutamate dehydrogenase reac-tion

L-Glutamate dehydrogenase occupies a central position in nitrogen metabolism

① Release of ammonia by hepatic L-glutmate dehydrogenase (GDH) (Fig. 28-8)

>> Conversion of α-amino nitrogen to ammonia by concerted action of Glu aminotransferase and GDH  transdeamination

>> Liver GDH is allosterically inhibited by ATP, GTP, and NADH and activated by ADP

GDH

Figure 28-8 The L-glutamate dehydrogenase reac-tion

α-amino nitrogen of AAs

Figure 27-1 The glutamate dehydrogenase reac-tion

α-amino nitrogen of AAs

Glutamate

- Reductive amidation of α-ketoglutarate by

glu-tamate dehydrogenase

- 1

_{step in the biosynthesis of the}

‘glutamate family’

1

Figure 28-9 Oxidative deamination catalyzed by L-amino acid oxi-dase

* Amino acid oxidases also remove nitrogen as ammonia

① L-amino acid oxidases : amino acids to an α-imino acid 

α-keto acid + ammonium ion (Fig. 28-9)

>>> Reduced flavin is reoxidized by oxygen  H₂O₂

Ammonia intoxification is life-threatening

>> Ammonia produced by enteric bacteria and absorbed into portal venous blood and produced by tissues  removed from circulation by liver and converted to urea

>> Ammonia is toxic to CNS; 10-20 μg/dL in peripheral blood >> Portal blood bypass the liver 

Ammonia intoxification: tremor, slurred speech, coma, death

Ammonia in brain+ α-ketoglutarate  Glu

: depletion of α-ketoglutarate impairs TCA cycle in neurons

GDH

1

Glutamine synthetase fixes ammonia as glutamine

>> Key function of Gln : Sequester ammonia in a nontoxic form

Figure 28-10 The glutamate synthetase reaction strongly fa-vors

glutamine synthesis

1

Glutaminase & asparginase deamidate glutamine & aspargine

① Hydrolytic release of ammonia from amide nitrogen of Gln by glutaminase (Fig. 28-11)

Asparagine

Asparagi-nase

Aspar-tate

Figure 28-11 The glutaminase reaction proceeds essentially

irreversibly

Glutaminase & asparginase deamidate glutamine & aspargine

>>> Hydrolytic release of ammonia from amide nitrogen of Gln by glutaminase (Fig. 28-11)

② Concerted action of Gln synthetase and glutaminase

 interconversion of free ammonium ion and Gln

Figure 28-11

Figure 28-10

Concerted reaction of aminotransferase and GDH

Formation & secretion of ammonia maintains acid-base balance

① Excretion into urine of ammonia

 cation conservation and regulation of acid-base balance ② Ammonia production from renal amino acid (esp. Gln):

* Urea is the major end product of nitrogen catabolism in humans

① Synthesis of 1 mol urea : 3 mol ATP + 1 mol ammonium ion + 1 mol of amino nitrogen of Aspartate ② Five enzymes (Fig. 28-12).

- Of the six participating AAs, N-acetylglutamate solely functions as enzyme activator

Mitochon-dria

Figure 28-12 Reactions and intermediates of urea biosyn-thesis

(1) Carbamoyl phosphate synthetase I initiates urea biosynthesis

by mito. carbamoyl phosphate synthase I (CPS I)

(cf. CPS II, cytosolic form of CPS1, use Gln as a source of nitrogen donor in pyrimidine biosynthesis (see Chap. 33)

② CPSI is active only in the presence of N-acetylglutamate (enzyme activator)

③ Concerted action of GDH and CPSI: nitrogen  carbamoyl phosphate

Mitochon-dria

(2) Carbamoyl phosphate plus ornithine forms citrulline

① L-Ornithine transcarbamoylase (mitochondria)  citrulline ② Entry of ornithine into mito. and exodus of citrulline from mito.: inner membrane transport system

Mitochon-dria

(3) Citrulline plus aspartate forms arginosuccinate

① Arginosuccinate synthetase links Asp and citrulline via the amino group of Asp and provides 2nd _{nitrogen of urea}

 Arginosuccinate

Mitochon-dria

(4) Cleavage of argininosuccinate forms arginine & fumarate

① Cleavage of argininosuccinate by argininosuccinase  arginine + fumarate (  malate  oxaloaetate) ② Transamination of oxaloacetate  Asp

Mitochon-dria

Cytosol

(5) Cleavage of arginine releases

urea

& re-forms ornithine

② Ornithine and lysine are inhibitors of arginase

③ Arginine as a precursor of NO by NO synthase (see Fig. 48-12)

Mitochon-dria

urea

Carbamoyl phosphate synthase I (CPS I)

is the pacemaker enzyme of the urea cycle

>>> Of the

six participating AAs

,

N-acetylglutamate

solely functions

as enzyme activator

Mitochon-dria

CPS I

① Metabolic disorders of urea biosynthesis:

② Defects in urea synthesis (severe in 1, 2) ->

ammonia intoxification

1) Hyperammonemia Type 1

- Carbamoyl phosphate synthase I

deficiency-> frequency (1:62,000)

2) Hyperammonemia type 2

① Deficiency of ornithine transcarbamoylase

(reaction 2)  X-chromosome linked

② Gln levels are elevated in blood, CSF, and

urine (enhanced Gln synthesis)

3) Citrullinemia

① High citrulline levels and excretion 

argininosuccinate synthetase

② Feeding arginine (  citrulline) or

ben-zoate  ammonia to hippurate via

glycine

4) Argininosuccinicaciduria

① Argininosuccinase (reaction 4)

② Feeding arginine and benzoate -> nitrogen

excretion

5) Hyperargininemia

-Elevated blood and CSF arginine levels

Metabolic fate of nonessential AAs

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