22.2: Biosynthesis of Amino Acids

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Learning Goals 

(Learning goals written by Claude, Sonnet 4.6, Anthropic)

Amino Acid Biosynthesis from Glycolytic and TCA Intermediates

• Identify the central metabolic precursor for each amino acid biosynthetic family — glucose-6-phosphate (histidine via the ribose phosphate pathway), 3-phosphoglycerate (serine, glycine, cysteine), phosphoenolpyruvate (aromatic amino acids via chorismate), pyruvate (alanine by transamination, valine/leucine/isoleucine via branched-chain pathways), α-ketoglutarate (glutamate, glutamine, proline, arginine via urea cycle intermediates), and oxaloacetate (aspartate, asparagine, methionine, threonine, lysine via the DAP or AAA pathways) — and explain how this organization reflects the direct connection between central metabolism and nitrogen incorporation.
• Explain why the aromatic amino acids (phenylalanine, tyrosine, tryptophan) are essential in humans but not in bacteria and plants — tracing their shared biosynthesis from phosphoenolpyruvate and erythrose-4-phosphate through the shikimate pathway to chorismate, from which separate branches produce prephenate (→ Phe/Tyr) or anthranilate (→ Trp) — and note that chorismate is also the precursor of folate and ubiquinone, making the shikimate pathway a high-value antibiotic and herbicide target.
• Describe the two pathways for lysine biosynthesis — the diaminopimelic acid (DAP) pathway (aspartate + pyruvate → diaminopimelic acid → lysine, used by bacteria, plants, and some fungi) versus the aminoadipic acid (AAA) pathway (α-ketoglutarate + acetyl-CoA → α-aminoadipate → lysine, used by fungi) — and explain why lysine is an essential amino acid in humans, who lack both pathways.

Cysteine, Glutathione, and Redox Homeostasis

• Explain the unique biochemical properties of cysteine that make it simultaneously essential for redox homeostasis and cytotoxic at elevated concentrations — describing how free cysteine reduces Fe³⁺ to Fe²⁺ (which drives the Fenton reaction generating ·OH radicals), undergoes auto-oxidation with O₂ to form cystine and H₂O₂, and disrupts cellular redox balance — and explain why the liver maintains low free cysteine by rapidly converting it to the less reactive glutathione and taurine.
• Describe glutathione (γ-glutamylcysteinylglycine, GSH) as the primary cellular antioxidant and redox buffer — explaining how the unusual γ-glutamyl linkage protects it from proteolytic degradation (enabling millimolar cellular concentrations versus the micromolar range for free cysteine), how glutathione peroxidase uses GSH to reduce H₂O₂ and lipid peroxides, how glutathione reductase uses NADPH to regenerate the reduced form, and how GSH depletion — triggered by blocking cysteine synthesis through CBS/CSE inhibition combined with BSO-mediated GCS inhibition — causes mitochondrial dysfunction, citric acid cycle enzyme damage, increased ROS from Complexes I and III, and ultimately systemic metabolic failure with dramatic body weight loss in animal models.

Introduction

By the time many students reach the study of amino acid biosynthesis, they have encountered numerous pathways, and learning new pathways for the amino acids may seem daunting, even though they can be clustered into subpathways. Most students are aware that, from a nutritional perspective, amino acids can be categorized into two types: nonessential and essential (those that require external dietary supplementation). These are shown for humans below.

• Nonessential amino acids: Alanine, Asparagine, Aspartate, Cysteine, Glutamate, Glutamine, Glycine, Proline, Serine, Tyrosine
• Essential amino acids: Arginine*, Histidine, Isoleucine, Leucine, Lysine, Methionine*, Phenylalanine*, Threonine, Tryptophan, Valine

Three of the essential amino acids can be made in humans, but need significant supplementation. Arginine is depleted during urea cycle processing. When cysteine levels are low, methionine is used to replace them, causing their levels to fall. If tyrosine is low, phenylalanine is used to replace it.

The amino acids can be synthesized from glycolytic and citric acid cycle intermediates as shown in Figure \(\PageIndex{1}\)

Figure \(\PageIndex{1}\): Summary amino acid synthesis from glycolytic and TCA intermediates

For this chapter subsection, we will provide only the basic synthetic pathways in abbreviated form, without delving into mechanistic or structural details (likely to the relief of readers and authors alike).

Amino acid synthesis from glycolytic intermediates

From Glucose-6-Phosphate: Histidine

The synthesis of histidine from a phosphorylated form of ribose (derived from glucose-6-phosphate) is shown in Figure \(\PageIndex{2}\).

Figure \(\PageIndex{2}\): Synthesis of histidine from a phosphorylated form of ribose

From 3-phosphoglycerate: Serine, Glycine, and Cysteine

The synthesis of serine, glycine, and cysteine from 3-phosphoglycerate is shown in Figure \(\PageIndex{3}\).

Figure \(\PageIndex{3}\): The synthesis of serine, glycine, and cysteine from 3-phosphoglycerate

Cysteine and control of body weight

Recent studies have shown that inhibiting cysteine synthesis results in significant weight loss. The figure below shows simplified pathways for Cys synthesis (blue) and consumption (reversible, green; and irreversible, brown). 

Figure: Simplified pathways for Cys synthesis (blue) and consumption (reversible, green; and irreversible, brown). Cystathionine β-synthase (CBS) and cystathionine γ-lyase (CSE) are involved in the trans-sulfuration pathway for cysteine synthesis. Varghese, A., Gusarov, I., Gamallo-Lana, B. et al. Unraveling cysteine-deficiency-associated rapid weight loss. Nature (2025). https://doi.org/10.1038/s41586-025-08996-y.  Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.  http://creativecommons.org/licenses/by-nc-nd/4.0/.

Dramatically decreasing cysteine leads to the depletion of glutathione (γ-glutamylcysteinylglycine, GSH), the main reducing agent (and antioxidant) in the cell, and CoASH, which is needed for the citric acid cycle and oxidation of fatty acids. Deletions of the mouse gene for CSE (red cross) and the addition of the inhibitor BSO (purple) decrease GSH synthesis, which proceeds through VNN (vanin/pantetheinase), GT (glutamyl transferase, and GCS (glutamate–cysteine ligase). Cysteine is a nonessential amino acid, but can be made essential through the inhibitions described in the above figure. These inhibitions led to a 30% reduction in mouse body weight in just one week.  White adipose tissue was lost and replaced with brown adipose tissue.  Accompanying these changes were large changes in transcription and mitochondrial dysfunction. Ultimately, this results in malfunctions in both the cell's integrated stress response (ISR) and oxidative stress response (OSR), inhibiting mitochondrial function.  Glycolytic and citric acid intermediates are lost in the urine, leading to weight loss.

What is so special about cysteine?  It is cytotoxic at high concentrations because it alters redox balance.  Hence, it has one of the lowest concentrations of free amino acids in the cell (typically 50–200 μM).  As you know from the study of protein structure, cysteine, as a free thiol, is a redox reagent (like β-mercaptoethanol or βME). βME is used at a concentration of 5% (about 7 M) in SDS-PAGE reducing sample preparation solutions.  At these very high levels, it reduces disulfide bonds in proteins by forming mixed disulfides, and then, upon further reaction with more βME, it forms free thiols.  Hence, it acts as a reducing agent.  At lower levels, it can form a persistent mixed disulfide with a cysteine or cystine in a target protein.  It is an oxidizing agent under these conditions (typical of a cellular environment). 

Given standard reduction potentials, cysteine can reduce Fe³⁺ to Fe²⁺, as shown below:

\begin{equation}
2 \mathrm{Cys}-\mathrm{SH}+2 \mathrm{Fe}^{3+} \longrightarrow \mathrm{RSSR}+2 \mathrm{Fe}^{2+}+2 \mathrm{H}^{+}
\end{equation}

The free Fe²⁺ is toxic to cells, in part by generating the hydroxyl free radical through the Fenton Reaction, as shown below:

\begin{equation}
\mathrm{Fe}^{2+}+\mathrm{H}_2 \mathrm{O}_2 \longrightarrow \mathrm{Fe}^{3+}+\mathrm{OH}^{-}+\cdot \mathrm{OH}
\end{equation}

This set of reactions becomes autocatalytic as the Fe³⁺ produced can re-enter the cycle. The hydroxyl radical oxidizes proteins, lipids, and nucleic acids and can also generate additional ROS.   

It can also react with O₂ in an autooxidation reaction, yielding cystine and ROS such as H₂O₂.  

\begin{equation}
2 \mathrm{Cys}-\mathrm{SH}+\mathrm{O}_2 \longrightarrow \mathrm{Cys}-\mathrm{S}-\mathrm{S}-\mathrm{Cys}+\mathrm{H}_2 \mathrm{O}_2
\end{equation}

Normally, ingested cysteine is kept low by liver uptake and conversion to GSH and taurine, which are less toxic. On low-cysteine diets, free intracellular cysteine is depleted as it is pulled into protein synthesis and used to maintain pools of CoASH and GSH.  The inhibitions described above amplify the dysfunction and lead to the results described. 

What's so special about GSH?  It is the main mitochondrial antioxidant. It is much more abundant in cells (0.5–15 mM). The γ-glutamyl group helps protect this tripeptide from proteolytic cleavage.  This helps contribute to its higher cellular concentration.  An enzyme, glutathione reductase, helps keep glutathione in its reduced form and at higher concentrations, acting as a redox buffering agent in cells. It is also much less reactive in the Fenton reaction. 

As shown below, GSH directly scavenges hydrogen peroxide (H₂O₂) and other peroxides using glutathione peroxidase.

\begin{equation}
2 \mathrm{GSH}+\mathrm{H}_2 \mathrm{O}_2 \xrightarrow{\text { glutathione peroxidase }} \mathrm{GSSG}+2 \mathrm{H}_2 \mathrm{O}
\end{equation}

and

\begin{equation}
2 \mathrm{GSH}+\mathrm{ROOH} \xrightarrow{\text { glutathione peroxidase }} \mathrm{GSSG}+\mathrm{ROH}+\mathrm{H}_2 \mathrm{O}
\end{equation}

When GSH is depleted, increased reactive oxygen species (ROS) produced at Complex I and III inhibit their activity through oxidative damage. Further damage to mitochondrial proteins reduces electron flow and ultimately decreases ATP production. Key mitochondrial citric acid cycle enzymes are also damaged.   

 

From Phosphoenolpyruvate: The Aromatics - Trp, Phe, and Tyr

The synthesis of the first of the biosynthetic pathways for the aromatic amino acids phenylalanine, tryptophan, and tyrosine from phosphoenolpyruvate up to chorismate is shown in Figure \(\PageIndex{4}\).

Figure \(\PageIndex{4}\): Synthesis of the first of the biosynthetic pathways for the aromatic amino acids phenylalanine, tryptophan, and tyrosine from phosphoenolpyruvate up to chorismate

Chorismate to tryptophan

The synthesis of the second half of the biosynthetic pathway for tryptophan from chorismate is shown in Figure \(\PageIndex{5}\)

Figure (\PageIndex{5}\): Synthesis of the second half of the biosynthetic pathways for the aromatic amino acid tryptophan from chorismate

Chorismate to Phe and Tyr

The synthesis of the second half of the biosynthetic pathway for phenylalanine and tyrosine from chorismate is shown in Figure \(\PageIndex{6}\)

Figure \(\PageIndex{6}\): Synthesis of the second half of the biosynthetic pathway for phenylalanine and tyrosine from chorismate

From Pyruvate: Ala, Val, Leu, Ile

Alanine can be easily synthesized from the alpha-keto acid pyruvate through a transamination reaction; therefore, we will focus our attention on the other branched-chain nonpolar amino acids, Val, Leu, and Ile.

The synthesis of valine, leucine, and isoleucine from pyruvate is shown in Figure \(\PageIndex{7}\).

Figure \(\PageIndex{7}\): The synthesis of valine, leucine, and isoleucine from pyruvate

TCA Intermediates

From α-ketoglutarate: Glu, Gln, Pro, Arg

Since amino acid metabolism is so complex, it is essential to continually review past learning. Figure \(\PageIndex{8}\) from section 18.2 shows the relationship among Glu, Gln, and keto acids.

Figure \(\PageIndex{8}\): Glutamate and glutamine synthesis from α-ketoglutarate

As shown in the figure, glutamic acid can be synthesized directly by transamination of α-ketoglutarate with an ammonia donor. In contrast, glutamine can be made by the action of glutamine synthase on glutamic acid.

Arginine is synthesized in the urea cycle, as previously discussed. It can be made from α-ketoglutarate through the following sequential intermediates: N-acetylglutamate, N-acetylglutamate-phosphate, N-acetylglutamate-semialdehyde, N-acetylornithine to N-acetylcitruline. It is deacetylated and enters the urea cycle.

The pathway for conversion of α-ketoglutarate to proline is shown in Figure \(\PageIndex{9}\).

 

Figure \(\PageIndex{9}\): Conversion of α-ketoglutarate to proline

From Oxaloacetate: Asp, Asn, Met, Thr, Lys

OAA to Aspartic Acid

This is a simple transamination

Aspartic Acid to Asparagine

This is catalyzed by the enzyme Asparagine Synthase, as shown in the reaction equation below:

Aspartate + Glutamine + ATP + H₂O → Asparagine + Glutamic Acids + AMP + PP_i

Aspartic Acid to Lysine

There are two pathways.

• The diaminopimelic acid (DAP) pathway utilizes aspartate and pyruvate, forming diaminopimelic acid as an intermediate. It is found in bacteria, some fungi, and archaea, as well as in plants.
• The aminoadipic acid (AAA) pathway utilizes α-ketoglutarate and acetyl-CoA to form aminoadipic acid as an intermediate. Fungi use it.

Here, we present the synthesis of lysine from aspartate and pyruvate using the diaminopimelic acid (DAP) pathway. The pathway is shown in Figure \(\PageIndex{10}\).

Figure \(\PageIndex{10}\): The synthesis of lysine from aspartic acid in the diaminopimelic acid DAP pathway

.

Aspartic acid to Threonine

The conversion of aspartic acid to threonine is shown in Figure \(\PageIndex{11}\).

Figure \(\PageIndex{11}\): The conversion of aspartic acid to threonine

Aspartic acid to Methionine

The conversion of aspartic acid to methionine is shown in Figure \(\PageIndex{12}\).

Figure \(\PageIndex{12}\): The conversion of aspartic acid to methionine

This SUMMARY GRAPHIC From Reactome shows "Cellular metabolism of amino acids and related molecules includes the pathways for the catabolism of amino acids, the biosynthesis of the nonessential amino acids (alanine, arginine, aspartate, asparagine, cysteine, glutamate, glutamine, glycine, proline, and serine) and selenocysteine, the synthesis of urea, and the metabolism of carnitine, creatine, choline, polyamides, melanin, and amine-derived hormones. The metabolism of amino acids provides a balanced supply for protein synthesis. In the fasting state, the catabolism of amino acids derived from the breakdown of skeletal muscle protein and other sources is coupled to the processes of gluconeogenesis and ketogenesis to meet the body’s energy needs in the absence of dietary energy sources."

Provided by Reactome. Citation Accessed on Wed, May 15, 2024.  Fabregat A, Sidiropoulos K, Viteri G, Marin-Garcia P, Ping P, Stein L, D'Eustachio P, Hermjakob H. Reactome diagram viewer: data structures and strategies to boost performance. Bioinformatics (Oxford, England). 2018 Apr;34(7) 1208-1214. doi: 10.1093/bioinformatics/btx752. PubMed PMID: 29186351. PubMed Central PMCID: PMC6030826.   Image:  https://reactome.org/PathwayBrowser/#/R-HSA-71291&PATH=R-HSA-1430728

Summary

(Summary written by Claude, Sonnet 4.6, Anthropic)

This chapter surveys the biosynthetic pathways for all 20 amino acids, organizing them by their central metabolic precursors and emphasizing the integration of amino acid synthesis with glycolysis and the TCA cycle, before examining in depth the special metabolic and physiological roles of cysteine and glutathione in cellular redox homeostasis.

Amino acids are classified nutritionally as nonessential (synthesizable by humans: Ala, Asn, Asp, Cys, Glu, Gln, Gly, Pro, Ser, Tyr) or essential (requiring dietary intake: His, Ile, Leu, Lys, Met, Phe, Thr, Trp, Val, and conditionally Arg). This distinction reflects which organisms possess the relevant biosynthetic enzymes. All biosynthetic pathways derive from a small number of central metabolic branch points, making amino acid biosynthesis an illustration of metabolic economy.

From glucose-6-phosphate (via the pentose phosphate pathway), histidine is synthesized through a multistep pathway using 5-phosphoribosyl-1-pyrophosphate (PRPP) as the carbon donor, with ATP providing both the nitrogen and an imidazole ring carbon — uniquely deriving its entire carbon skeleton from the ribose phosphate rather than from the α-keto acid precursors used by most other amino acids. From 3-phosphoglycerate, serine is produced by oxidation, transamination, and phosphatase steps; glycine arises from serine via serine hydroxymethyltransferase (SHMT, transferring one carbon to FH4); and cysteine is synthesized by replacement of serine's hydroxyl group with sulfide derived from homocysteine via the transsulfuration pathway (CBS and CSE enzymes), using methionine-derived SAM as the ultimate sulfur source.

From phosphoenolpyruvate and erythrose-4-phosphate (from the pentose phosphate pathway), the shikimate pathway builds the central intermediate chorismate in seven steps, providing the branch point for all three aromatic amino acids. From chorismate, a two-step pathway through prephenate produces phenylalanine (by dehydratase) and tyrosine (by dehydrogenase); humans can additionally convert phenylalanine to tyrosine by phenylalanine hydroxylase (BH4-dependent), making tyrosine conditionally essential. A separate branch from chorismate through anthranilate and indole-3-glycerol-phosphate produces tryptophan in five additional steps. Because the shikimate pathway is absent in animals but essential in bacteria, fungi, and plants, its enzymes are important targets for antibiotics (such as fosfomycin) and herbicides (glyphosate inhibits EPSP synthase). From pyruvate, alanine is produced by a single transamination reaction with glutamate; valine and leucine share a four-step pathway from two pyruvate molecules; and isoleucine synthesis begins with threonine deamination and then merges with the valine pathway using α-ketobutyrate.

From α-ketoglutarate (a TCA cycle intermediate), glutamate is produced directly by reductive amination or transamination, and serves as the amino group donor for most other transamination reactions; glutamine is produced from glutamate by glutamine synthetase (consuming one ATP); proline is produced by reduction of glutamate through glutamate-5-semialdehyde and cyclization; and arginine is synthesized through N-acetylglutamate intermediates before entering the urea cycle. From oxaloacetate, aspartate is produced by transamination and is the common precursor for asparagine (via asparagine synthase using glutamine as nitrogen donor, consuming ATP), methionine (via β-aspartyl intermediates including homoserine, cystathionine, and homocysteine), threonine (via phosphoaspartate and homoserine), and lysine (via the DAP pathway in bacteria and plants using diaminopimelic acid as a key intermediate, or the AAA pathway using α-ketoglutarate and acetyl-CoA in fungi).

The chapter devotes particular attention to the exceptional biochemistry of cysteine and glutathione, supported by recent experimental findings of dramatic physiological consequences when cysteine is depleted. Free cysteine is maintained at unusually low cellular concentrations (50–200 μM) because it is cytotoxic at higher levels: its free thiol reduces Fe³⁺ to Fe²⁺ (driving the Fenton reaction to produce ·OH radicals), undergoes auto-oxidation with O₂ to form cystine and H₂O₂, and can form persistent mixed disulfides with protein cysteines, disrupting signaling and structure. Normal cysteine catabolism in the liver — rapid conversion to glutathione and taurine — prevents accumulation while preserving the sulfur for biologically essential functions.

Glutathione (γ-Glu-Cys-Gly, GSH) is the primary intracellular antioxidant, present at 0.5–15 mM in most cells. Its γ-glutamyl peptide bond (connecting the γ-carboxylate of glutamate to cysteine's amine, rather than the standard α-carboxylate) protects it from most cellular proteases, explaining why it accumulates to millimolar concentrations while free cysteine remains micromolar. GSH directly reduces H₂O₂ and lipid peroxides via glutathione peroxidase (generating GSSG), and the reduced form is regenerated by NADPH-dependent glutathione reductase, maintaining the cell's redox buffering capacity. GSH is also the main mitochondrial antioxidant; when depleted, ROS produced at Complexes I and III accumulate and oxidatively damage mitochondrial proteins, reducing electron transport chain activity and ATP production while also impairing TCA cycle enzymes. Recent animal studies strikingly illustrate the indispensability of cysteine: combined inhibition of cysteine biosynthesis (deletion of the CSE gene) and GSH synthesis (BSO inhibition of GCS) produced 30% body weight loss in mice within one week — accompanied by loss of white adipose tissue, replacement with brown adipose tissue, transcriptional reprogramming, mitochondrial dysfunction, impaired integrated stress response (ISR) and oxidative stress response (OSR), and loss of glycolytic and TCA cycle intermediates in urine. These results reveal that, although nominally nonessential in its dietary classification, cysteine becomes physiologically essential when its synthetic pathways or GSH-mediated utilization are blocked, underscoring the interconnection among amino acid metabolism, antioxidant defense, and systemic metabolic integrity.