GO:0140773 NAD-dependent protein demyristoylase activity

NAD-dependent protein demyristoylase activity (GO:0140773) is a molecular function that catalyzes the removal of a myristoyl group from a modified lysine residue on a target protein, using NAD+ as an essential co-substrate. This activity belongs to the sirtuin class of enzymes, which couple deacylation to NAD+ hydrolysis and produce nicotinamide and an acyl-ADP-ribose intermediate. The reaction is chemically defined as N6-tetradecanoyl-L-lysyl-[protein] + NAD+ + H2O = tetradecanoyl-ADP-ribose + L-lysyl-[protein] + nicotinamide, and it represents a key regulatory mechanism for dynamic protein lipidation. Researchers are increasingly interested in demyristoylation because it reverses a hydrophobic modification that controls protein localization, stability, and interactions. Unlike irreversible lipid modifications, demyristoylation allows cells to rapidly remodel signaling complexes in response to metabolic and stress cues. The discovery that SIRT2 efficiently removes myristoyl groups from peptide substrates established this activity as a bona fide enzymatic function rather than a secondary side reaction. More recently, SIRT6 was shown to demyristoylate ATF2, linking this activity directly to vascular endothelial barrier protection through the PRKCD/VE-cadherin pathway. For biomedical researchers, GO:0140773 provides a precise annotation for experiments involving fatty-acylation dynamics, sirtuin biology, and NAD+ metabolism. Understanding which enzymes carry this activity, which substrates they target, and how the reaction is regulated is essential for interpreting proteomic and functional data in cardiovascular, metabolic, and inflammatory disease models.

In simple terms, GO:0140773 describes an enzyme that acts like a molecular eraser: it removes a 14-carbon fatty acid called myristate from a lysine residue on a protein. The reaction requires NAD+ and water, and it produces three products: the demyristoylated protein, nicotinamide, and tetradecanoyl-ADP-ribose. This definition is based on the QuickGO entry for GO:0140773 and reflects the catalytic chemistry shared by NAD+-dependent sirtuin deacylases.

GO:0140773 matters because it defines a reversible, NAD+-dependent mechanism for controlling protein lipidation, a process that influences membrane targeting, protein-protein interactions, and signal transduction. The activity connects cellular energy status, via NAD+ availability, to the dynamic regulation of fatty-acylated proteins, making it a focal point for research on metabolic stress, inflammation, and vascular biology. Because sirtuins such as SIRT2 and SIRT6 can remove long-chain acyl groups, demyristoylation provides a biochemical explanation for how cells fine-tune signaling beyond classical acetylation.

The following genes and proteins are directly implicated in NAD-dependent protein demyristoylase activity or its physiological consequences, based on the verified literature.

NAD-dependent protein demyristoylase activity is regulated primarily by the availability of its obligate co-substrate NAD+, which fluctuates with cellular metabolic state and stress. Because the reaction consumes NAD+ and produces nicotinamide, product inhibition by nicotinamide can modulate enzyme turnover, a feature well documented for sirtuin deacylases. In the SIRT6-ATF2 axis, demyristoylation is embedded in a signaling cascade that controls PRKCD and VE-cadherin, indicating that upstream stress and inflammatory signals can influence the functional output of this activity. Structural determinants, including the hydrophobic acyl-binding pocket, also regulate substrate preference and catalytic efficiency.

SIRT6 lysine-demyristoylates ATF2 to ameliorate vascular injury via the PRKCD/VE-cadherin pathway, which regulates the vascular endothelial barrier. This places GO:0140773 at the intersection of protein deacylation and endothelial barrier integrity, suggesting that loss of demyristoylase activity could contribute to vascular leak and injury. Experimental models of endothelial barrier disruption are therefore relevant for studying this activity.

Because demyristoylation requires NAD+, conditions that alter NAD+ metabolism, such as metabolic stress and inflammation, can indirectly affect this activity. ATF2 is a stress-responsive transcription factor, and its demyristoylation by SIRT6 links the modification to stress signaling pathways. This connection supports investigating GO:0140773 in metabolic and inflammatory disease contexts.

SIRT2 exhibits efficient demyristoylase activity, and sirtuins are widely studied in aging, neurodegeneration, and cancer. Although direct disease links for demyristoylation specifically remain an active area, the enzymatic properties of SIRT2 provide a foundation for exploring how this activity contributes to sirtuin-associated pathologies. Structural and kinetic data on SIRT2 demyristoylation can guide hypothesis-driven disease research.

Kinetic studies using synthetic myristoylated peptides allow direct measurement of demyristoylase catalytic efficiency and substrate specificity. These assays typically monitor NAD+ consumption or product formation, such as nicotinamide or tetradecanoyl-ADP-ribose. SIRT2 demyristoylase activity was established using such kinetic approaches.

X-ray crystallography and structural analysis of sirtuins reveal the hydrophobic acyl-binding pocket that accommodates the myristoyl chain. Structural studies of SIRT2 provided mechanistic insight into why long-chain acyl groups are efficiently removed. These methods help distinguish demyristoylases from deacetylases.

Endothelial barrier integrity can be assessed using permeability assays and junction protein localization in cells expressing wild-type or mutant SIRT6 and ATF2. The SIRT6-ATF2-PRKCD-VE-cadherin axis provides a defined pathway for functional readouts. These assays connect molecular demyristoylation to physiological barrier function.

Detection of myristoylated versus demyristoylated proteins can be approached through modification-specific enrichment and mass spectrometry, guided by the known chemistry of GO:0140773. Comparing wild-type and enzyme-deficient cells helps identify candidate substrates. Such workflows are essential for mapping the substrate landscape of demyristoylases.

Researchers studying NAD-dependent protein demyristoylase activity-related genes often need to determine whether a candidate gene is causally involved in a specific phenotype, such as endothelial barrier protection or metabolic stress responses. EDITGENE provides CRISPR-based cell model services that enable precise genetic perturbations for such mechanistic studies.

NAD-dependent protein demyristoylase activity (GO:0140773) is a chemically defined, NAD+-consuming enzymatic function that removes myristate from lysine residues on proteins. SIRT2 and SIRT6 are the best-characterized enzymes with this activity, and the SIRT6-ATF2-PRKCD-VE-cadherin axis provides a concrete physiological example linking demyristoylation to vascular endothelial barrier protection. As a molecular function annotation, GO:0140773 enables precise interpretation of experiments involving protein lipidation, sirtuin biology, and NAD+ metabolism. Future research using CRISPR-based models, structural biology, and modification-specific proteomics will help define the full substrate repertoire and disease relevance of demyristoylation. EDITGENE supports these efforts with customizable cell model and screening services tailored to demyristoylase research.