GO:0004372 glycine hydroxymethyltransferase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0004372 describes the enzymatic activity that reversibly converts glycine plus (6R)-5,10-methylene-5,6,7,8-tetrahydrofolate and water into L-serine and (6S)-5,6,7,8-tetrahydrofolate, linking one-carbon metabolism to serine and glycine homeostasis.
• The major human enzymes carrying this activity are the cytosolic and mitochondrial serine hydroxymethyltransferases SHMT1 and SHMT2, with SHMT2 being especially important in mitochondria and in tumors.
• SHMT2 activity can be reversed under specific metabolic conditions, driving glycine depletion and contributing to acetaminophen hepatotoxicity in metabolic dysfunction-associated steatotic liver disease.
• SHMT2 is regulated by phosphorylation, succinylation and transcriptional programs, and its dysregulation supports antioxidant defence, purine synthesis and oncogenesis in lung, thyroid and other cancers.
• Loss of PYCR2 increases cerebral glycine via SHMT2, linking this activity to neurodegeneration and to mitochondrial folate-dependent translation.
• CRISPR knockout, point-mutation, knock-in and overexpression models are essential to dissect SHMT1/SHMT2-dependent one-carbon flux and to validate therapeutic hypotheses.
Description
Glycine hydroxymethyltransferase activity (GO:0004372) is a molecular function that catalyzes the reversible interconversion of glycine and serine using tetrahydrofolate derivatives as one-carbon carriers. This activity sits at the intersection of amino acid metabolism, one-carbon metabolism and folate cycling, and it is essential for supplying methyl groups for nucleotide synthesis, methylation reactions and mitochondrial translation. In humans, the enzymes SHMT1 and SHMT2 are the principal carriers of this activity, with SHMT2 localized to mitochondria and SHMT1 predominantly cytosolic. Because the reaction is reversible, its direction depends on substrate availability and compartment-specific metabolic demands, making it a central node in cancer, liver disease and neurodevelopmental disorders. Researchers study GO:0004372 to understand how cells balance serine, glycine and one-carbon units, and to identify vulnerabilities in tumors and metabolic diseases.
glycine hydroxymethyltransferase activity At A Glance
| GO ID | GO:0004372 |
|---|---|
| GO term | glycine hydroxymethyltransferase activity |
| Ontology | molecular_function |
| Synonym | serine hydroxymethyltransferase activity; serine aldolase activity; L-serine hydroxymethyltransferase activity; 5,10-methylenetetrahydrofolate:glycine hydroxymethyltransferase activity |
| Major function | Reversible conversion of glycine and (6R)-5,10-methylene-tetrahydrofolate to L-serine and (6S)-5,6,7,8-tetrahydrofolate |
| Major enzymes | SHMT1 (cytosolic), SHMT2 (mitochondrial) |
| Cofactor | Pyridoxal phosphate (PLP) dependent |
| Pathways | One-carbon metabolism, folate cycle, serine-glycine interconversion, mitochondrial translation |
| Disease relevance | Cancer, MASLD/acetaminophen hepatotoxicity, neurodegeneration, metabolic disorders |
What Is GO:0004372?
GO:0004372, glycine hydroxymethyltransferase activity, is defined by QuickGO as the catalysis of the reaction: (6R)-5,10-methylene-5,6,7,8-tetrahydrofolate + glycine + H2O = (6S)-5,6,7,8-tetrahydrofolate + L-serine. In other words, the enzyme transfers a hydroxymethyl group from a folate derivative to glycine to produce serine, or runs the reverse reaction to generate glycine and 5,10-methylene-tetrahydrofolate. This activity is synonymous with serine hydroxymethyltransferase activity, serine aldolase activity and related names, and it is a molecular_function term in the Gene Ontology.
Why Is glycine hydroxymethyltransferase activity Important in Cell Biology?
GO:0004372 is important because it controls the balance between serine and glycine and feeds one-carbon units into folate-dependent processes such as purine synthesis, methylation and mitochondrial translation. In cancer, SHMT2 activity supports antioxidant defence and oncogenesis, and its inhibition or reversal can alter tumor growth and therapy response. In liver disease, reversed SHMT2 activity drives glycine depletion and acetaminophen hepatotoxicity in MASLD. In the brain, loss of PYCR2 increases glycine via SHMT2, linking this activity to neurodegeneration. Thus, understanding GO:0004372 is essential for metabolic research, drug discovery and CRISPR-based disease modeling.
• Provides serine for protein synthesis and one-carbon units for nucleotide synthesis.
• Supports mitochondrial translation through folate-dependent tRNA methylation.
• Enables antioxidant defence in KRAS-mutant lung cancer with LKB1/KEAP1 loss.
• Is regulated by phosphorylation and m6A modification in lung adenocarcinoma.
• Can reverse direction to deplete glycine in MASLD and acetaminophen toxicity.
• Links to succinate-mediated control of purine synthesis via succinylation.
• Contributes to cerebral glycine accumulation in PYCR2 deficiency.
• Is a targetable vulnerability in persister cancer cells via H4K20me3 repression.
• Serves as a biomarker and therapeutic target in undifferentiated thyroid cancer.
• Enables CRISPR screens to identify metabolic dependencies in tumors.
Molecular Mechanism of glycine hydroxymethyltransferase activity
Substrate binding and PLP-dependent catalysis
In simple terms: The enzyme uses a vitamin B6-derived cofactor to move a chemical group between folate and glycine.
Glycine hydroxymethyltransferase activity requires pyridoxal phosphate (PLP) as a cofactor. The enzyme binds glycine and (6R)-5,10-methylene-5,6,7,8-tetrahydrofolate, and catalyzes the transfer of a hydroxymethyl group to glycine, producing L-serine and (6S)-5,6,7,8-tetrahydrofolate. This reaction is reversible, and the direction depends on substrate availability and compartment-specific metabolic needs.
Compartmentalization: SHMT1 and SHMT2
In simple terms: Two main enzymes do this job in different parts of the cell.
In humans, SHMT1 is predominantly cytosolic, while SHMT2 is mitochondrial. SHMT2 is especially important for mitochondrial one-carbon metabolism and for supplying glycine and serine in tumors. Multi-omics analyses in undifferentiated thyroid cancer highlight the mitochondrial one-carbon pathway involving SHMT2.
Reversed activity and glycine depletion
In simple terms: Sometimes the enzyme runs backward and consumes glycine instead of making it.
Under specific conditions, SHMT2 can operate in reverse, depleting glycine and contributing to acetaminophen hepatotoxicity in MASLD. This reversed activity is a key example of how GO:0004372 directionality can influence disease outcomes.
Regulation by phosphorylation and succinylation
In simple terms: Chemical tags on the enzyme can switch its activity up or down.
SHMT2 is regulated by phosphorylation, which affects oncogenesis through m6A modification in lung adenocarcinoma. Succinate accumulation suppresses de novo purine synthesis through succinylation-mediated control of the mitochondrial folate cycle, implicating SHMT2 regulation. These post-translational modifications fine-tune GO:0004372 activity in response to metabolic signals.
Role in mitochondrial translation and antioxidant defence
In simple terms: The enzyme helps mitochondria make proteins and fight oxidative stress.
Mitochondrial translation requires folate-dependent tRNA methylation, a process linked to one-carbon metabolism and SHMT2. Concurrent loss of LKB1 and KEAP1 enhances SHMT-mediated antioxidant defence in KRAS-mutant lung cancer, showing how GO:0004372 supports redox balance.
Key Genes Involved in GO:0004372 glycine hydroxymethyltransferase activity
The following genes and proteins are directly or functionally linked to glycine hydroxymethyltransferase activity (GO:0004372) based on the verified literature.
| Gene | Major Role | Research Relevance |
|---|---|---|
| SHMT1 | Cytosolic serine hydroxymethyltransferase | Provides one-carbon units for nucleotide synthesis and methylation |
| SHMT2 | Mitochondrial serine hydroxymethyltransferase | Central to mitochondrial one-carbon metabolism, cancer and liver disease |
| MTHFD2 | Mitochondrial folate cycle enzyme | Supports one-carbon flux and purine synthesis |
| MTHFD1L | Mitochondrial folate cycle enzyme | Contributes to formate production for purine synthesis |
| PYCR2 | Proline synthesis enzyme | Loss increases cerebral glycine via SHMT2 |
| LKB1 (STK11) | Tumor suppressor kinase | Loss enhances SHMT-mediated antioxidant defence |
| KEAP1 | Oxidative stress regulator | Loss with LKB1 enhances SHMT dependency |
| KRAS | Oncogene | Mutant KRAS context shows SHMT vulnerability |
| MTHFR | Folate metabolism enzyme | Generates 5-methyl-THF for methionine synthesis |
| MTR | Methionine synthase | Links folate cycle to methylation |
| GCSH | Glycine cleavage system H protein | Interacts with glycine metabolism |
| GLDC | Glycine decarboxylase | Contributes to glycine homeostasis |
| AMT | Glycine cleavage system T protein | Supports glycine cleavage |
| MTHFD1 | Cytosolic folate cycle enzyme | Supports one-carbon metabolism |
| ATIC | Purine synthesis enzyme | Downstream of one-carbon flux |
| GART | Purine synthesis enzyme | Downstream of one-carbon flux |
| PPAT | Purine synthesis enzyme | Downstream of one-carbon flux |
How Is glycine hydroxymethyltransferase activity Regulated?
Glycine hydroxymethyltransferase activity is regulated at multiple levels. SHMT2 phosphorylation modulates oncogenesis through m6A modification in lung adenocarcinoma. Succinate accumulation suppresses de novo purine synthesis through succinylation-mediated control of the mitochondrial folate cycle, affecting SHMT2 function. Loss of LKB1 and KEAP1 enhances SHMT-mediated antioxidant defence, indicating metabolic stress pathways regulate this activity. In MASLD, reversed SHMT2 activity is driven by metabolic conditions, leading to glycine depletion. Additionally, mitochondrial translation requires folate-dependent tRNA methylation, linking one-carbon status to SHMT2 regulation.
glycine hydroxymethyltransferase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| SHMT2 | MASLD and acetaminophen hepatotoxicity | SHMT2 knockout or point-mutation hepatocytes |
| SHMT2 | Lung adenocarcinoma oncogenesis | SHMT2 phosphorylation-site knock-in in lung cancer cells |
| SHMT2 | Undifferentiated thyroid cancer | SHMT2 overexpression or knockout in thyroid cancer models |
| SHMT2 | KRAS-mutant lung cancer antioxidant defence | LKB1/KEAP1 double knockout with SHMT2 knockout |
| PYCR2 | Neurodegeneration with cerebral glycine accumulation | PYCR2 knockout neurons with SHMT2 knockdown |
Cancer metabolism and oncogenesis
SHMT2 activity supports oncogenesis in lung adenocarcinoma through phosphorylation and m6A modification. In KRAS-mutant lung cancer with LKB1 and KEAP1 loss, SHMT-mediated antioxidant defence is enhanced, promoting tumor survival. Undifferentiated thyroid cancer shows upregulation of the mitochondrial one-carbon pathway involving SHMT2. Persister cancer cells exhibit H4K20me3-mediated repression of inflammatory genes, representing a targetable vulnerability linked to one-carbon metabolism.
Liver disease and acetaminophen hepatotoxicity
In metabolic dysfunction-associated steatotic liver disease (MASLD), reversed SHMT2 activity drives glycine depletion and acetaminophen hepatotoxicity. This highlights GO:0004372 as a potential therapeutic target in liver disease.
Neurodegeneration and cerebral glycine
Loss of PYCR2 causes neurodegeneration by increasing cerebral glycine levels via SHMT2. This connects glycine hydroxymethyltransferase activity to neurodevelopmental and neurodegenerative disorders.
Mitochondrial translation and metabolic disorders
Mitochondrial translation requires folate-dependent tRNA methylation, a process dependent on one-carbon metabolism and SHMT2. Succinate accumulation suppresses purine synthesis through succinylation-mediated control of the mitochondrial folate cycle, linking GO:0004372 to metabolic disorders.
From glycine hydroxymethyltransferase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does SHMT2 loss alter glycine and serine levels? | SHMT2 knockout cell lines |
| Does SHMT2 phosphorylation affect oncogenesis? | Phospho-mutant knock-in in lung cancer cells |
| Does reversed SHMT2 activity drive hepatotoxicity? | SHMT2 point-mutation or knockout hepatocytes |
| Does SHMT2 support antioxidant defence? | SHMT2 overexpression in KRAS-mutant lung cancer cells |
| Does PYCR2 loss increase glycine via SHMT2? | PYCR2 knockout with SHMT2 knockdown |
| Does succinylation regulate folate cycle? | SHMT2 succinylation-site mutant knock-in |
How to Study the glycine hydroxymethyltransferase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Metabolomics | Serine, glycine and one-carbon metabolites | Quantify SHMT2 activity |
| Stable isotope tracing | Flux through one-carbon metabolism | Determine reaction direction |
| CRISPR knockout screens | Gene dependencies | Identify regulators of GO:0004372 |
| Phosphoproteomics | SHMT2 phosphorylation | Study oncogenic signaling |
| Succinylome analysis | SHMT2 succinylation | Link succinate to folate cycle |
| Mitochondrial translation assay | Folate-dependent tRNA methylation | Assess mitochondrial function |
| Multi-omics integration | Pathway upregulation | Thyroid cancer analysis |
| H4K20me3 ChIP-seq | Inflammatory gene repression | Persister cancer cells |
Metabolomics and flux analysis
Metabolomics and stable isotope tracing measure serine, glycine and one-carbon metabolites to quantify GO:0004372 activity. These methods are used to assess SHMT2 directionality and pathway flux in cancer and liver disease models.
CRISPR screens and functional genomics
CRISPR knockout screens identify genes that modulate glycine hydroxymethyltransferase activity and its downstream pathways. These screens are applied to discover metabolic vulnerabilities in tumors.
Proteomics and post-translational modification analysis
Proteomics detects phosphorylation and succinylation of SHMT2, linking modifications to activity changes. This is used to study regulation of GO:0004372 in cancer and metabolic stress.
Imaging and mitochondrial translation assays
Imaging and mitochondrial translation assays assess folate-dependent tRNA methylation and mitochondrial function. These methods connect GO:0004372 to mitochondrial translation and metabolic disorders.
How CRISPR Can Be Used to Study GO:0004372 glycine hydroxymethyltransferase activity
Knockout
CRISPR knockout of SHMT1 or SHMT2 is used to eliminate glycine hydroxymethyltransferase activity and assess metabolic and phenotypic consequences. Knockout models help determine whether a gene is causally involved in serine-glycine homeostasis and disease.
Point Mutation
Point mutations in SHMT2, such as phosphorylation-site mutants, allow researchers to dissect how specific residues regulate activity and oncogenesis. These models are used to test whether post-translational modifications drive disease phenotypes.
Knock-in
Knock-in of tagged or mutant SHMT2 enables tracking of protein localization, interaction and function in cells. Knock-in models are valuable for studying succinylation or other modifications.
Overexpression
Overexpression of SHMT2 or SHMT1 is used to enhance glycine hydroxymethyltransferase activity and test its role in antioxidant defence and tumor growth. Overexpression models help validate gain-of-function hypotheses.
How EDITGENE Supports glycine hydroxymethyltransferase activity Research
Researchers studying glycine hydroxymethyltransferase activity-related genes often need to determine whether a candidate gene is causally involved in metabolic and disease phenotypes. EDITGENE provides CRISPR-based cell model services to enable such functional validation.
Contact EDITGENE today to design your custom CRISPR model for glycine hydroxymethyltransferase activity research.
Frequently Asked Questions About glycine hydroxymethyltransferase activity
What is glycine hydroxymethyltransferase activity?
It is the enzymatic activity defined by GO:0004372 that reversibly converts glycine and (6R)-5,10-methylene-5,6,7,8-tetrahydrofolate to L-serine and (6S)-5,6,7,8-tetrahydrofolate.
What genes are involved in glycine hydroxymethyltransferase activity?
The main human genes are SHMT1 and SHMT2, which encode cytosolic and mitochondrial serine hydroxymethyltransferases.
What is the GO ID for glycine hydroxymethyltransferase activity?
The GO ID is GO:0004372.
How is glycine hydroxymethyltransferase activity regulated?
It is regulated by phosphorylation, succinylation and metabolic signals such as succinate accumulation.
What diseases are linked to glycine hydroxymethyltransferase activity?
It is linked to cancer, MASLD and acetaminophen hepatotoxicity, neurodegeneration and metabolic disorders.
What is the role of SHMT2 in cancer?
SHMT2 supports oncogenesis, antioxidant defence and mitochondrial one-carbon metabolism in lung and thyroid cancers.
Can glycine hydroxymethyltransferase activity run in reverse?
Yes, reversed SHMT2 activity can deplete glycine and drive acetaminophen hepatotoxicity in MASLD.
How do researchers study glycine hydroxymethyltransferase activity?
They use metabolomics, stable isotope tracing, CRISPR screens, proteomics and mitochondrial translation assays.
What CRISPR models are used for SHMT2 research?
Knockout, point-mutation, knock-in and overexpression models are used to dissect SHMT2 function.
Why is glycine hydroxymethyltransferase activity important for mitochondrial translation?
It supports folate-dependent tRNA methylation, which is required for mitochondrial translation.
Conclusion
Glycine hydroxymethyltransferase activity (GO:0004372) is a central molecular function in one-carbon metabolism, controlling serine-glycine balance and supplying methyl groups for nucleotide synthesis, methylation and mitochondrial translation. Its dysregulation is implicated in cancer, liver disease and neurodegeneration, making it a key target for metabolic research. CRISPR-based models are indispensable for dissecting the causal roles of SHMT1, SHMT2 and related genes, and for developing therapeutic strategies.
References
- 1. Ghrayeb A et al.. 2024. Serine synthesis via reversed SHMT2 activity drives glycine depletion and acetaminophen hepatotoxicity in MASLD.. Cell Metab 36(1):116-129.e7 PMID: 38171331
- 2. Han T et al.. 2024. Phosphorylated SHMT2 Regulates Oncogenesis Through m(6)A Modification in Lung Adenocarcinoma.. Adv Sci (Weinh) 11(18):e2307834 PMID: 38460155
- 3. Lee SE et al.. 2024. Unraveling the role of the mitochondrial one-carbon pathway in undifferentiated thyroid cancer by multi-omics analyses.. Nat Commun 15(1):1163 PMID: 38331894
- 4. Lee HM et al.. 2024. Concurrent loss of LKB1 and KEAP1 enhances SHMT-mediated antioxidant defence in KRAS-mutant lung cancer.. Nat Metab 6(7):1310-1328 PMID: 38877143
- 5. Nengroo MA et al.. 2025. Accumulation of succinate suppresses de novo purine synthesis through succinylation-mediated control of the mitochondrial folate cycle.. Mol Cell 85(22):4215-4228.e9 PMID: 41161310
- 6. Morscher RJ et al.. 2018. Mitochondrial translation requires folate-dependent tRNA methylation.. Nature 554(7690):128-132 PMID: 29364879
- 7. Escande-Beillard N et al.. 2020. Loss of PYCR2 Causes Neurodegeneration by Increasing Cerebral Glycine Levels via SHMT2.. Neuron 107(1):82-94.e6 PMID: 32330411
- 8. Ramponi V et al.. 2025. H4K20me3-Mediated Repression of Inflammatory Genes Is a Characteristic and Targetable Vulnerability of Persister Cancer Cells.. Cancer Res 85(1):32-51 PMID: 39476057
