GO:0047961 glycine N-acyltransferase activity: Mechanism, Genes and Research Methods
Research-grade guide for scientists and biopharma professionals
Key Takeaways
• GO:0047961 glycine N-acyltransferase activity is a molecular_function defined as the catalysis of the reaction acyl-CoA + glycine = CoA + N-acylglycine.
• The reaction is a glycine conjugation step that converts xenobiotic and endogenous acyl-CoAs into more water-soluble N-acylglycines for excretion.
• Human GLYAT is the principal enzyme for glycine conjugation of benzoyl-CoA and other acyl-CoAs, and its catalytic mechanism has been dissected by mutagenesis and kinetics.
• GLYATL2 is a related transferase that produces long-chain N-acyl glycines, and its activity is regulated by reversible lysine acetylation.
• Biallelic loss-of-function variants in GLYAT cause a novel inborn error of metabolism with abnormal acylglycine profiles, and common GLYAT variants influence interindividual glycine conjugation capacity.
• CRISPR knockout, point-mutation, knock-in and overexpression models enable causal testing of GLYAT and GLYATL2 variants in glycine conjugation and drug metabolism.
Description
Glycine N-acyltransferase activity (GO:0047961) is a molecular_function that catalyzes the transfer of an acyl group from an acyl-CoA donor to the amino group of glycine, yielding coenzyme A and an N-acylglycine. This reaction is the terminal step of glycine conjugation, a major phase II detoxification route for aromatic and other carboxylic acids in humans and other mammals. The reaction is chemically simple but physiologically important because it converts hydrophobic acyl-CoAs into more polar, excretable N-acylglycines such as hippurate. Researchers study GO:0047961 to understand xenobiotic metabolism, mitochondrial acyl-CoA handling, and inherited disorders of glycine conjugation. The human genome encodes several glycine N-acyltransferase enzymes, including GLYAT and GLYATL2, which differ in substrate preference and regulation. Because glycine conjugation capacity varies widely between individuals, functional characterization of GLYAT variants is directly relevant to drug metabolism and personalized medicine.
glycine N-acyltransferase activity At A Glance
| GO ID | GO:0047961 |
|---|---|
| GO term | glycine N-acyltransferase activity |
| Ontology | molecular_function |
| Definition | Catalysis of the reaction: acyl-CoA + glycine = CoA + N-acylglycine |
| Synonyms | acyl-CoA:glycine N-acyltransferase activity; glycine acyltransferase activity; glycine-N-acylase activity |
| Major function | Transfer of acyl groups from acyl-CoA donors to glycine, producing N-acylglycines and free CoA |
| Representative human genes | GLYAT, GLYATL2 |
| Pathway context | Glycine conjugation / phase II xenobiotic metabolism |
| Disease relevance | Inborn error of metabolism due to GLYAT deficiency; interindividual variation in drug conjugation |
What Is GO:0047961?
In the Gene Ontology, GO:0047961 glycine N-acyltransferase activity is defined as catalysis of the reaction acyl-CoA + glycine = CoA + N-acylglycine. In other words, the enzyme takes an activated acyl group carried by coenzyme A and attaches it to the amino group of glycine, releasing free CoA and producing an N-acylglycine. The term is a molecular_function and is synonymous with acyl-CoA:glycine N-acyltransferase activity, glycine acyltransferase activity, and glycine-N-acylase activity. The reaction is a glycine conjugation reaction that increases the water solubility of the acyl group and supports its excretion.
Why Is glycine N-acyltransferase activity Important in Cell Biology?
GO:0047961 is important because glycine conjugation is a conserved detoxification and metabolic pathway that determines how efficiently the body clears aromatic carboxylic acids, drug metabolites, and endogenous acyl-CoAs. The reaction catalyzed by glycine N-acyltransferase enzymes directly controls the formation of hippurate and other N-acylglycines, which are measurable biomarkers of mitochondrial and hepatic acyl-CoA metabolism. Loss of GLYAT function causes a newly described inborn error of metabolism with abnormal acylglycine excretion, and common GLYAT variants contribute to interindividual differences in glycine conjugation capacity. In addition, GLYATL2-derived N-acyl glycines have been implicated in signaling, and their production is tuned by lysine acetylation. Therefore, this GO term sits at the intersection of enzymology, drug metabolism, inherited disease, and cell signaling.
• Defines the terminal step of glycine conjugation, a major phase II detoxification route.
• Produces hippurate and other N-acylglycines used as clinical and metabolic biomarkers.
• GLYAT deficiency is a novel inborn error of metabolism with a distinct biochemical phenotype.
• Common GLYAT sequence variants alter glycine conjugation capacity and drug handling.
• GLYATL2 generates long-chain N-acyl glycines with potential signaling roles.
• GLYATL2 activity is regulated by reversible lysine acetylation, linking metabolism to acetylation status.
• Kinetic and mutagenesis studies have defined the catalytic residues and mechanism of human GLYAT.
• Functional characterization of GLYAT variants supports personalized prediction of conjugation capacity.
• The pathway is relevant to acetaminophen and other drug metabolism models in vivo.
• CRISPR models allow causal testing of GLYAT and GLYATL2 variants in isogenic backgrounds.
Molecular Mechanism of glycine N-acyltransferase activity
Substrate recognition and acyl-CoA binding
In simple terms: The enzyme first grabs an activated acyl group that is carried by coenzyme A.
Glycine N-acyltransferase enzymes bind acyl-CoA substrates and position the reactive thioester for nucleophilic attack by glycine. Human GLYAT accepts benzoyl-CoA and related aromatic acyl-CoAs, consistent with its role in hippurate synthesis. GLYATL2 prefers longer-chain acyl-CoAs and produces long-chain N-acyl glycines, indicating that substrate specificity is enzyme-specific. The acyl-CoA binding step is therefore a key determinant of which N-acylglycine products are formed in a given cell or tissue.
Catalytic transfer to glycine
In simple terms: The acyl group is handed from coenzyme A to the amino group of glycine.
The catalytic reaction follows the GO definition acyl-CoA + glycine = CoA + N-acylglycine, in which glycine acts as the acyl acceptor and coenzyme A is released. Mutagenesis and kinetic studies of human GLYAT have identified residues that contribute to catalysis and substrate positioning, supporting a direct acyl-transfer mechanism. The reaction does not require ATP because the acyl-CoA thioester already provides the activated acyl group. Product release yields free CoA and an N-acylglycine, completing the conjugation step.
Enzyme variants and catalytic efficiency
In simple terms: Small changes in the enzyme sequence can change how fast the reaction runs.
Functional characterization of three human GLYAT variants showed that sequence changes can alter glycine conjugation to benzoyl-CoA, providing direct evidence that catalytic efficiency is genotype-dependent. Frequent sequence variants of human GLYAT have been catalogued in the context of inborn errors of metabolism, reinforcing the link between genotype and conjugation capacity. A homozygous nonsense variant in GLYAT causes glycine N-acyltransferase deficiency, demonstrating that loss of catalytic activity produces a measurable metabolic phenotype. These observations make GLYAT a model for connecting enzyme variants to metabolic flux.
Regulation by acetylation and cellular context
In simple terms: The activity of related enzymes can be switched up or down by chemical tags added to the protein.
Human GLYATL2 activity is regulated by reversible lysine acetylation, which modulates the production of glycine-conjugated signaling molecules. This post-translational control links glycine N-acyltransferase activity to the broader cellular acetylation state. In vivo, Xiaoyao powder attenuated acetaminophen-induced liver injury in association with upregulation of GLYAT, indicating that GLYAT expression and activity can be modulated by interventions. Together, these findings show that GO:0047961 is not a fixed flux but is tuned by both genetic variation and regulatory inputs.
Key Genes Involved in GO:0047961 glycine N-acyltransferase activity
The following genes and proteins are directly or functionally associated with glycine N-acyltransferase activity (GO:0047961) and its physiological context.
| Gene | Major Role | Research Relevance |
|---|---|---|
| GLYAT | Principal human glycine N-acyltransferase for aromatic acyl-CoAs such as benzoyl-CoA | Variant functional characterization and deficiency modeling |
| GLYATL2 | Transferase that produces long-chain N-acyl glycines in humans | Substrate specificity and acetylation-dependent regulation |
| GLYATL1 | Glycine N-acyltransferase-like family member | Comparative enzymology of the GLYAT family |
| GLYATL3 | Glycine N-acyltransferase-like family member | Family-level substrate and expression comparisons |
| ACSM2A | Generates acyl-CoA substrates from carboxylic acids | Upstream supply of acyl-CoA for glycine conjugation |
| ACSM2B | Mitochondrial acyl-CoA synthetase contributing to acyl-CoA pools | Substrate availability for GLYAT reactions |
| ACSM3 | Acyl-CoA synthetase involved in short-chain acyl-CoA metabolism | Metabolic context of glycine conjugation |
| GLYCTK | Glycine metabolism enzyme influencing glycine availability | Glycine donor pool for N-acylglycine synthesis |
| SLC25A10 | Mitochondrial dicarboxylate carrier linked to acyl-CoA metabolism | Mitochondrial substrate transport context |
| SLC25A1 | Mitochondrial citrate carrier influencing acetyl-CoA pools | Indirect metabolic context for acyl-CoA supply |
| BAAT | Bile acid-CoA:amino acid N-acyltransferase, related conjugation enzyme | Comparative N-acyltransferase mechanism |
| NAT1 | Arylamine N-acetyltransferase, another phase II conjugation enzyme | Comparative phase II metabolism context |
| NAT2 | Arylamine N-acetyltransferase with polymorphic activity | Interindividual variation comparison |
| UGT1A1 | Glucuronidation enzyme, alternative conjugation route | Pathway competition and detoxification context |
| SULT1A1 | Sulfotransferase, alternative phase II conjugation enzyme | Comparative conjugation capacity |
| CYP2E1 | Cytochrome P450 generating reactive acyl metabolites | Acetaminophen metabolism context |
| NQO1 | Antioxidant enzyme in drug-induced liver injury models | Liver injury model readout |
How Is glycine N-acyltransferase activity Regulated?
Glycine N-acyltransferase activity is regulated at multiple levels. At the post-translational level, human GLYATL2 is controlled by reversible lysine acetylation, which modulates its ability to produce glycine-conjugated signaling molecules. At the genetic level, frequent sequence variants in GLYAT influence glycine conjugation capacity and are relevant to inborn errors of metabolism, and functional characterization of GLYAT variants shows that catalytic efficiency toward benzoyl-CoA can differ between alleles. At the physiological level, glycine availability and acyl-CoA supply shape flux through the reaction, and factors influencing interindividual variation in glycine conjugation have been reviewed. In vivo, GLYAT expression can be upregulated in the context of attenuated acetaminophen-induced liver injury, indicating that the pathway responds to pharmacological and dietary interventions.
glycine N-acyltransferase activity and Human Disease
| Gene | Disease / Biology | Potential Experimental Model |
|---|---|---|
| GLYAT | Glycine N-acyltransferase deficiency, a novel inborn error of metabolism | Patient-derived cells or GLYAT knockout hepatocyte-like cells |
| GLYAT | Interindividual variation in glycine conjugation and drug metabolism | Isogenic cell lines carrying GLYAT point mutations |
| GLYAT | Acetaminophen-induced liver injury and detoxification | Mouse or cell models with GLYAT overexpression or knockout |
| GLYATL2 | Production of long-chain N-acyl glycine signaling molecules | GLYATL2 knockout and acetylation-site mutant cells |
| GLYAT family | Comparative enzymology of N-acyltransferases | Overexpression of GLYAT, GLYATL1, GLYATL2 and GLYATL3 in cell lines |
GLYAT deficiency as an inborn error of metabolism
A homozygous nonsense variant in GLYAT was identified in a patient with glycine N-acyltransferase deficiency, establishing this enzyme defect as a novel inborn error of metabolism. The biochemical hallmark is an abnormal acylglycine profile reflecting impaired glycine conjugation. This finding directly links loss of GO:0047961 activity to a human Mendelian disease and provides a benchmark for variant interpretation.
Interindividual variation in drug and xenobiotic conjugation
Common GLYAT sequence variants are frequent in human populations and have been discussed in the context of inborn errors of metabolism and variable conjugation capacity. Functional studies of GLYAT variants demonstrate that amino acid changes can alter glycine conjugation to benzoyl-CoA, which is relevant to how individuals handle carboxylic acid drugs and metabolites. Reviews of glycine conjugation emphasize that interindividual variation in this pathway has pharmacological and toxicological consequences.
Acetaminophen-induced liver injury and metabolic stress
In an experimental model of acetaminophen-induced liver injury, Xiaoyao powder attenuated injury in association with modulation of gut microbiota and upregulation of GLYAT. This connects glycine N-acyltransferase activity to hepatic stress responses and detoxification of drug-derived acyl metabolites. The study supports the use of GLYAT expression and activity as readouts in liver injury models.
N-acyl glycine signaling and metabolic regulation
GLYATL2 produces long-chain N-acyl glycines, and its activity is regulated by reversible lysine acetylation, implicating these products in signaling processes. Dysregulation of such signaling molecules could contribute to metabolic phenotypes, although the disease associations remain an active area of research. This makes GLYATL2 an attractive target for studies linking conjugation chemistry to cell signaling.
From glycine N-acyltransferase activity-Related Genes to Experimental Models
| Research Question | Suitable Model |
|---|---|
| Does loss of GLYAT abolish glycine conjugation of benzoyl-CoA? | GLYAT knockout cell line or organoid |
| Does a specific GLYAT missense variant reduce catalytic efficiency? | Point-mutation knock-in of the variant into an isogenic background |
| Can a patient GLYAT variant be corrected to restore activity? | Knock-in of wild-type GLYAT or base-edited correction |
| Where is GLYAT localized and how is it expressed? | Tagged knock-in with fluorescent or affinity tag |
| Does GLYATL2 acetylation change N-acyl glycine output? | Overexpression of wild-type and acetylation-site mutants |
| Does increased GLYAT protect against drug-induced liver injury? | GLYAT overexpression in hepatocyte models |
How to Study the glycine N-acyltransferase activity Process
| Method | What It Measures | Typical Application |
|---|---|---|
| Enzyme activity assay with acyl-CoA and glycine | Formation of CoA or N-acylglycine | Variant functional characterization |
| Targeted metabolomics | N-acylglycine and hippurate levels | Diagnosis and pathway flux |
| Sanger or next-generation sequencing | GLYAT and GLYATL2 variants | Inborn error and variant discovery |
| Recombinant protein kinetics | Catalytic efficiency and substrate specificity | Mechanistic enzymology |
| Western blot and qPCR | GLYAT expression levels | Intervention and liver injury studies |
| Acetylation-site mutagenesis | Effect of lysine acetylation on GLYATL2 | Post-translational regulation |
| CRISPR knockout and knock-in | Causal role of genes and variants | Isogenic functional genomics |
| Overexpression in cell models | Gain-of-function effects on conjugation | Protection and signaling studies |
Enzymatic assays for glycine N-acyltransferase activity
Direct measurement of GO:0047961 uses acyl-CoA and glycine as substrates and detects CoA or N-acylglycine formation. Kinetic characterization of recombinant GLYAT and its variants allows determination of catalytic efficiency and substrate preference. Such assays are the primary way to confirm that a variant alters enzyme function.
Metabolomics and acylglycine profiling
Because the products of the reaction are N-acylglycines, targeted metabolomics can quantify hippurate and related species in cells, plasma, or urine. Abnormal acylglycine profiles were central to identifying GLYAT deficiency as an inborn error of metabolism. Metabolite profiling also provides a functional readout in liver injury and intervention studies.
Genotyping and variant functional characterization
Sequencing of GLYAT and related genes identifies common and rare variants that may affect glycine conjugation. Functional characterization of variants in recombinant systems or isogenic cell lines links genotype to enzyme activity. This approach supports variant classification in inborn errors of metabolism.
Expression and regulation studies
Transcript and protein measurements of GLYAT and GLYATL2 reveal how expression responds to pharmacological or dietary interventions. Acetylation studies, including acetylation-site mutants, define post-translational control of GLYATL2 activity. Together with enzyme assays, these methods provide a multi-level view of pathway regulation.
How CRISPR Can Be Used to Study GO:0047961 glycine N-acyltransferase activity
Knockout
CRISPR knockout of GLYAT or GLYATL2 creates isogenic models to test whether loss of GO:0047961 activity abolishes specific N-acylglycine products. Such models are useful for validating the contribution of each enzyme to glycine conjugation and for reproducing the biochemical phenotype of GLYAT deficiency. Knockout cells can also be challenged with acyl-CoA-generating drugs to measure pathway flux.
Point Mutation
Point-mutation knock-in allows precise testing of GLYAT variants identified in patients or population studies without confounding background variation. Isogenic lines carrying a single missense or nonsense allele can be compared directly for enzyme activity and metabolite output. This approach is essential for classifying variants of uncertain significance in glycine conjugation disorders.
Knock-in
Knock-in of wild-type or tagged GLYAT enables expression under endogenous regulatory control and supports localization and interaction studies. Tagged knock-in lines can be used to measure protein abundance and turnover in response to interventions. Corrective knock-in or base editing can also test whether restoring GLYAT activity rescues the deficiency phenotype.
Overexpression
Overexpression of GLYAT or GLYATL2 provides gain-of-function models to test whether increased glycine N-acyltransferase activity alters metabolite profiles or protects against injury. Overexpression of acetylation-site mutants of GLYATL2 can dissect post-translational regulation of N-acyl glycine production. These models complement knockout studies by defining the upper range of pathway activity.
How EDITGENE Supports glycine N-acyltransferase activity Research
Researchers studying glycine N-acyltransferase activity-related genes often need to determine whether a candidate gene is causally involved in glycine conjugation, drug metabolism, or inherited metabolic disease. Establishing causality requires controlled genetic models in which GLYAT or GLYATL2 is removed, mutated, tagged, or overexpressed in an isogenic background. EDITGENE provides end-to-end CRISPR cell model generation and screening services to support such studies, from knockout and point-mutation lines to knock-in reporters and overexpression pools.
Contact EDITGENE today to design your custom CRISPR model for glycine N-acyltransferase activity research.
Frequently Asked Questions About glycine N-acyltransferase activity
What is glycine N-acyltransferase activity?
Glycine N-acyltransferase activity (GO:0047961) is a molecular_function that catalyzes the reaction acyl-CoA + glycine = CoA + N-acylglycine, transferring an acyl group from an acyl-CoA donor to glycine.
What genes are involved in glycine N-acyltransferase activity?
The main human genes are GLYAT, which conjugates aromatic acyl-CoAs such as benzoyl-CoA, and GLYATL2, which produces long-chain N-acyl glycines.
What is the GO definition of GO:0047961?
GO:0047961 is defined as catalysis of the reaction acyl-CoA + glycine = CoA + N-acylglycine, with synonyms including acyl-CoA:glycine N-acyltransferase activity and glycine-N-acylase activity.
What does glycine N-acyltransferase do in the body?
It performs the terminal step of glycine conjugation, converting acyl-CoAs into more water-soluble N-acylglycines such as hippurate for excretion.
Is GLYAT deficiency a disease?
Yes, a homozygous nonsense variant in GLYAT causes glycine N-acyltransferase deficiency, a novel inborn error of metabolism with abnormal acylglycine profiles.
How is glycine N-acyltransferase activity measured?
It is measured by enzyme assays using acyl-CoA and glycine as substrates, often combined with metabolomic detection of N-acylglycine products.
What is the difference between GLYAT and GLYATL2?
GLYAT primarily conjugates aromatic acyl-CoAs such as benzoyl-CoA, whereas GLYATL2 produces long-chain N-acyl glycines and is regulated by lysine acetylation.
Can GLYAT variants affect drug metabolism?
Yes, functional characterization shows that GLYAT variants can alter glycine conjugation to benzoyl-CoA, and common variants contribute to interindividual variation in conjugation capacity.
How do I make a GLYAT knockout cell line?
CRISPR knockout of GLYAT can be generated in relevant cell types to test loss of glycine conjugation and N-acylglycine production, and such models are available through EDITGENE.
Why is glycine conjugation important in toxicology?
Glycine conjugation increases the water solubility of carboxylic acids and their metabolites, supporting their excretion and contributing to detoxification of drugs such as acetaminophen.
Conclusion
Glycine N-acyltransferase activity (GO:0047961) is a well-defined molecular_function that catalyzes the transfer of acyl groups from acyl-CoA donors to glycine, producing N-acylglycines and free coenzyme A. Its human representatives, GLYAT and GLYATL2, connect this simple conjugation chemistry to drug metabolism, mitochondrial acyl-CoA handling, inherited metabolic disease, and N-acyl glycine signaling. The discovery of GLYAT deficiency as an inborn error of metabolism and the functional characterization of GLYAT variants underscore the clinical and pharmacological relevance of this activity. CRISPR-based knockout, point-mutation, knock-in, and overexpression models provide the causal tools needed to move from variant association to mechanism and to test therapeutic or dietary interventions that modulate glycine conjugation.
References
- 1. Nourbakhsh M et al.. 2025. Glycine N-Acyltransferase Deficiency due to a Homozygous Nonsense Variant in the GLYAT: A Novel Inborn Error of Metabolism.. JIMD Rep 66(5):e70032 PMID: 40747359
- 2. Xiong Y et al.. 2026. Xiaoyao powder attenuates acetaminophen-induced liver injury through modulating gut microbiota and upregulating GLYAT.. J Ethnopharmacol 359:121079 PMID: 41423158
- 3. Rohwer JM et al.. 2021. Functional Characterisation of Three Glycine N-Acyltransferase Variants and the Effect on Glycine Conjugation to Benzoyl-CoA.. Int J Mol Sci 22(6) PMID: 33803916
- 4. Waluk DP et al.. 2010. Identification of glycine N-acyltransferase-like 2 (GLYATL2) as a transferase that produces N-acyl glycines in humans.. FASEB J 24(8):2795-803 PMID: 20305126
- 5. Schulke D et al.. 2021. Frequent sequence variants of human glycine N-acyltransferase (GLYAT) and inborn errors of metabolism.. Biochimie 183:30-34 PMID: 33567294
- 6. van der Sluis R et al.. 2017. New insights into the catalytic mechanism of human glycine N-acyltransferase.. J Biochem Mol Toxicol 31(11) PMID: 28759163
- 7. Waluk DP et al.. 2012. Reversible lysine acetylation regulates activity of human glycine N-acyltransferase-like 2 (hGLYATL2): implications for production of glycine-conjugated signaling molecules.. J Biol Chem 287(20):16158-67 PMID: 22408254
- 8. Badenhorst CP et al.. 2013. Glycine conjugation: importance in metabolism, the role of glycine N-acyltransferase, and factors that influence interindividual variation.. Expert Opin Drug Metab Toxicol 9(9):1139-53 PMID: 23650932
