Acetylation Antibodies: How Do They Reveal the Key Role of N-terminal Acetylation in Autophagy Regulation?

Recent Advances

I. What Are the Unique Characteristics of N-terminal Acetylation Modification?

Protein N-terminal acetylation is one of the most prevalent post-translational modifications in eukaryotes, occurring with a frequency exceeding 80%. Unlike classical acetylation occurring on lysine side chains, N-terminal acetylation specifically refers to the transfer of an acetyl group from acetyl-CoA to the α-amino group at the protein N-terminus. This reaction is catalyzed by the N-terminal acetyltransferase (NAT) family. Among them, the Nat3 enzyme possesses unique substrate specificity due to its highly conserved GCN5-related acetyltransferase domain. Nat3 specifically recognizes protein sequences starting with methionine, followed by aspartic acid, glutamic acid, asparagine, or glutamine. This precise substrate selectivity provides important clues for its functional positioning within specific cellular pathways.
 
II. How Was Nat3 Identified as a Key Autophagy Regulator?

Through genome-wide yeast genetic screening, researchers first identified Nat3 as a core regulator of autophagy. In-depth cell biology studies revealed that Nat3 deletion completely blocks autophagic flux, with a phenotypic severity comparable to mutants of the core autophagy gene atg6. At the molecular level, in Nat3-deficient cells, the autophagy substrate Ape1 fails to enter the vacuole for degradation and instead forms abnormal aggregates in the cytoplasm. Simultaneously, the localization of the autophagosome marker protein ATG8 is significantly disrupted, changing from a normal punctate pattern to a diffuse distribution throughout the cell. Through elegantly designed functional complementation experiments, researchers further confirmed that only Nat3 with intact catalytic activity could restore autophagy function, indicating that its acetyltransferase activity is necessary for its role in autophagy regulation.
 
III. How to Validate Nat3's Specific Substrates?

Researchers employed a multi-technique strategy to systematically validate Nat3's specific substrates. First, bioinformatic predictions combined with proteomic analysis identified actin (Act1) and the GTPase Vps1 as candidate substrates. Subsequently, co-immunoprecipitation experiments using N-terminal acetylation-specific antibodies confirmed that both proteins indeed undergo N-terminal acetylation. To obtain more direct evidence, researchers further utilized high-resolution mass spectrometry to precisely identify the modification sites at the N-terminal methionine residues of these two proteins. This series of rigorous experiments not only confirmed Nat3's substrate specificity but also fully demonstrated the irreplaceable role of acetylation antibodies in studying protein post-translational modifications.

 
IV. How Does Vps1 Acetylation Affect the Autophagy Process?

Vps1, a dynamin-like GTPase, plays a key role in the late stages of autophagy. In-depth studies revealed that Vps1 participates in autophagy by regulating the membrane fusion between autophagosomes and the vacuole. When Vps1 undergoes N-terminal acetylation, its molecular conformation changes, significantly enhancing its interaction with the SNARE protein Vam3, thereby promoting the correct assembly and functional activation of the SNARE complex. To clarify the functional significance of the acetylation modification, researchers constructed mutants mimicking a constitutively acetylated state (D2M) and a non-acetylatable state (D2P). Functional analysis showed that only the acetylated form of Vps1 could effectively mediate autophagosome-vacuole fusion, whereas the non-acetylated form caused the autophagy process to be blocked at the final step.
 
V. What Role Does Act1 Acetylation Play in Autophagy?

During the initiation stage of autophagy, the N-terminal acetylation of Act1 is crucial for autophagosome formation. Research indicates that acetylation promotes the correct assembly and dynamic remodeling of actin filaments by modulating the charge properties and spatial conformation of Act1. These actin filaments provide the necessary track system for the directional transport of Atg9 vesicles, ensuring the efficient delivery of membrane components from the Golgi apparatus to the phagophore assembly site. Detailed cellular imaging analysis showed that in Nat3-deficient cells, the co-localization of Atg9 vesicles with the actin network was significantly reduced, and vesicle transport was severely impaired, directly preventing the normal formation of autophagosomes.

 
VI. How Does Nat3 Precisely Regulate Autophagy Through a Dual Mechanism?

Nat3 achieves precise regulation of autophagy by coordinating two key steps upstream and downstream. At the upstream regulatory level, Nat3 catalyzes the N-terminal acetylation of Act1, promoting the functional assembly of the actin cytoskeleton, which provides the structural basis for Atg9 vesicle transport and ensures normal autophagosome formation. At the downstream regulatory level, Nat3-mediated acetylation of Vps1 enhances its interaction with the SNARE complex, promoting the specific fusion of autophagosomes with the vacuole to complete the final steps of autophagy. The elucidation of this complete molecular pathway systematically clarifies, for the first time, the central role of N-terminal acetylation in the autophagy regulatory network, revealing the important function of protein post-translational modifications in cellular quality control.
 
VII. What is the Scientific Value and Application Prospect of This Research?

This research holds significant scientific importance at multiple levels. In basic research, it not only reveals a new function of N-terminal acetylation in cellular autophagy for the first time but also provides an innovative research paradigm for studying protein post-translational modifications. The successful application of acetylation antibodies demonstrates their unique value in detecting specific site modifications, offering an important technical reference for related fields. In translational medicine, this discovery provides a new theoretical framework for understanding the pathogenesis of diseases associated with autophagy abnormalities, including neurodegenerative diseases, metabolic diseases, and cancer. Based on the in-depth study of the Nat3 regulatory pathway, new strategies for modulating autophagy may be developed in the future, providing new targets and directions for treating related diseases.
 
VIII. Which Companies Supply Acetylation Antibodies?

Hangzhou Start Biotech Co., Ltd. has independently developed the "Acetylated Lysine Rabbit Polyclonal Antibody" (Product Name: Acetyllysine Rabbit polyclonal antibody, Product Code: S0B0655). This is a protein post-translational modification detection tool characterized by broad specificity, high affinity, and excellent stability. Prepared using carefully designed synthetic acetylated lysine peptide immunogens and rigorously validated across multiple platforms including Immunoprecipitation, Western Blot, and Immunofluorescence, it holds broad application value in fields such as epigenetic regulation, metabolic pathway research, and cell signal transduction.

Core Product Advantages:
* Broad Recognition Capability and High Specificity: This product accurately recognizes acetylation modifications on protein lysine residues and demonstrates exceptional detection capability for acetylation across various protein backgrounds. Cross-validation with modified/unmodified peptides shows extremely high specificity, providing a reliable tool for non-specific acetylation detection.
* Excellent Affinity and Batch Consistency: The product exhibits high affinity, effectively enriching and detecting endogenously acetylated proteins. Strict immune serum pool management and purification processes ensure high consistency in performance across different batches, providing stable support for long-term research projects.
* Suitable for Various Key Application Scenarios: This product is an ideal tool for the following research areas:
* Histone Acetylation Research: For detecting acetylation levels at sites like Histone H3 and H4, exploring their role in gene expression regulation.
* Non-Histone Acetylation Detection: For studying acetylation modifications and their functional regulation of non-histone proteins like transcription factors and metabolic enzymes.
* Epigenetic Drug Screening: For screening drugs like histone deacetylase inhibitors and evaluating their efficacy.
* Cell Metabolism and Signaling Pathway Research: For exploring the regulatory mechanisms of acetylation in cell metabolic reprogramming and signal transduction pathways.
* Professional Technical Support: We provide detailed product technical documentation, including specificity validation data, experimental protocols for various application platforms, and professional technical support, fully committed to assisting customers in achieving breakthroughs in the field of protein modification and function research. For more details about the "Acetylated Lysine Rabbit Polyclonal Antibody" (Product Code S0B0655) or to request a sample test, please feel free to contact us.
 
Product Information
 

Catalog No.	Product Name	Product Parameters	Size	Price
S0F0004	Anti-acetyllysine agarose Beads		1ml	Inquiry
S0B0878	Histone H4 (acetyl K5 + K8 + K12 + K16) Recombinant Rabbit mAb (S-902-37)	Host : Rabbit Conjugation : Unconjugated	100μl	$350
S0B0546	Histone H3 (acetyl K27) Recombinant Rabbit mAb (S-699-50)	Host : Rabbit Conjugation : Unconjugated	100μl	$350
S0B1154	Histone H3 (acetyl K18) Recombinant Rabbit mAb (S-1688-90)	Host : Rabbit Conjugation : Unconjugated	100μl	$350
S0B0755	Histone H3 (acetyl K14) Recombinant Rabbit mAb (S-R398)	Host : Rabbit Conjugation : Unconjugated	100μl	$350
S0B0655	Acetyllysine Rabbit polyclonal antibody	Host : Rabbit Conjugation : Unconjugated	100μl	$350