Creative Biolabs maintains a prominent standing within antibody research and development circles, specializing in post-translational modification (PTM)-targeting antibody production. Through the proprietary High-Affi™ platform, we offer fatty acylation-specific antibodies with enhanced binding capabilities to international clients.

Background

Fatty acylation refers to the covalent bonding of fatty acid groups to protein structures. This modification exhibits distinct characteristics compared to alternative hydrophobic alterations like prenylation or cholesterol modification, particularly through direct myristate or palmitate conjugation forming N-myristoylation and palmitoylation respectively.

Protein N-myristoylation represents a widespread biological mechanism involving irreversible myristate attachment to N-terminal glycine residues. Mediated through N-myristoyltransferase (NMT) activity, this modification occurs co-translationally and post-translationally across multiple biological kingdoms, spanning mammals, plants, fungi, protozoa, and viral entities. The process critically influences cellular navigation systems, membrane association dynamics, and molecular interaction networks, while also participating in apoptosis regulation, structural transformations, and viral particle assembly. Disrupted myristoylation patterns correlate with oncogenic progression and enhanced viral pathogenicity, particularly through NMT overexpression observed in multiple malignancies. Enhanced c-Src myristoylation, for instance, demonstrates capacity to stimulate vascular proliferation, neoplastic growth, and metastatic behavior, establishing this modification pathway as a candidate for pharmacological intervention.

Fig.1 Protein N-Myristoylation. Distributed under CC BY-SA 3.0, from Wiki, without modification.

Distinct from other lipidations, S-palmitoylation operates through reversible cysteine residue modification via 16-carbon fatty acid attachment. This dynamic process remains governed by counteractive enzymatic systems: palmitoyl acyltransferases (PATs) facilitate palmitate addition while thioesterases execute its removal. Ubiquitous across eukaryotic, prokaryotic, and viral biology, palmitoylation modulates molecular stability, structural conformation, intracellular routing, and partnership formation between biomolecules. Current research implicates palmitoylation dysregulation in neurodegenerative pathologies and multiple carcinogenic processes.

Fig.2 Protein S-palmitoylation.¹

Myristoylation and palmitoylation frequently demonstrate interdependent modification patterns on shared protein substrates. While solitary myristoylation permits transient membrane engagement through weaker associations, subsequent palmitoylation reinforces membrane anchoring through sustained hydrophobic interactions. Such dual modification mechanics prove particularly consequential in G protein-coupled receptor (GPCR) signaling cascades. These acylation events additionally interact with parallel modification systems, as evidenced by G protein signal transduction mechanisms: palmitoylated α subunits combine with prenylated γ subunits, while subsequent myristoylation enables plasma membrane adherence and receptor engagement.

Antibody Types

Polyclonal Antibodies: Created through animal immunization, these contain antibodies targeting multiple antigen regions. Effective for broad detection, though batch consistency varies.

Monoclonal Antibodies: Clonal B-cell derivatives with singular epitope specificity. Ideal for standardized, precise experimental applications.

Myristoylation Antibodies: Identify myristate-linked N-terminal glycines. Critical for tracking modified proteins in cellular processes and disease mechanisms.

Palmitoylation Antibodies: Detect cysteine-bound palmitate despite modification reversibility. Technically demanding but essential for studying dynamic protein roles in signaling and pathology.

Discovery Strategy

Monospecific Anti-Fatty Acylation Polyclonal Antibody Production

Generating polyclonal antibodies with fatty acylation specificity follows two critical phases. Initially, animals receive immunizations using carrier protein-coupled fatty acylated peptides to stimulate diverse antibody generation. Post-immunization, serum undergoes purification via affinity columns functionalized with matching acylated peptides, selectively capturing modification-specific antibodies while discarding irrelevant binders. This approach merges polyclonal antibodies' collective binding strength with refined specificity, yielding affordable reagents capable of detecting lipid-modified targets without compromising accuracy.

Phage Display Strategy for Anti-Fatty Acylation Monoclonal Antibody Discovery

Harnessing phage display technology, synthetic libraries are screened to discover monoclonal antibodies against fatty acylations. This approach can screen both immune libraries, derived from immunized animals for targeted responses, and prefabricated libraries from non-immune or human sources. The method involves iterative exposure of phage-displayed antibody fragments to acylated antigens, followed by enrichment and amplification of high-affinity binders, and sequencing, to reconstruct antibody genes for recombinant production. This in vitro system explores vast antibody diversity, enabling the targeting of challenging epitopes and transient modifications, with library sizes surpassing conventional immunology.

Hybridoma Strategy for Anti-Fatty Acylation Monoclonal Antibody Discovery

Hybridoma methodology remains a gold standard for monoclonal antibody development against fatty acylations. Immunized animals provide splenic B cells fused with myeloma counterparts, creating immortalized antibody factories. Post-fusion screening identifies clones producing acyl-specific antibodies, followed by subcloning to guarantee genetic uniformity. These cell lines enable perpetual antibody production with batch consistency. The technique's reliability stems from decades of optimization, yielding antibodies with exceptional affinity and stability while supporting industrial-grade manufacturing for clinical or research applications.

Product

Types of PTM

In addition to the fatty acylation-specific antibody, Creative Biolabs also provides a comprehensive list of PTM-specific antibody production services of your choice.

Service Highlights

Lipidated Hapten Expertise: We design haptens replicating fatty acylation structures (e.g., myristoylation, palmitoylation) to generate antibodies that selectively recognize modified proteins over unmodified counterparts.

High-Affi™ for Lipid Targets: Our High-Affi™ technology refines antibody selection for lipid modifications, yielding reagents with unmatched affinity and specificity for acylated targets.

Lipid-Dependent Binding Screens: We screen candidates using lipid competition assays and modified/unmodified peptide comparisons to isolate antibodies strictly dependent on fatty acyl groups.

Hydrophobic Antigen Handling: Specialized methods address hydrophobic antigen challenges, ensuring effective presentation of lipidated peptides/proteins for successful immunization.

Q&A

Q: What fatty acylation modifications can your antibodies target?

A: We develop antibodies against diverse fatty acylation modifications, covering common types like N-myristoylation and S-palmitoylation alongside other less common acylations. Our hapten customization options let us adapt precisely to your lipid modification requirements.

Q: How do you ensure the specificity of the generated antibodies for the fatty acylated protein?

A: Specificity assurance involves a three-tiered strategy. Initial screenings use modified/unmodified peptide pairs to eliminate cross-reactive clones. Intermediate validation employs truncated protein variants to confirm lipid dependency. For critical applications, we recommend supplementary testing through Western blot comparisons of wild-type versus modification-deficient cell lysates, ensuring antibodies exclusively recognize the acylated form in complex biological contexts.

Q: What are the advantages of your High-Affi™ platform for fatty acylation antibody discovery?

A: The High-Affi™ system integrates advanced mutagenesis protocols with lipid-specific binding analytics, driving affinity enhancements exceeding conventional methods. This platform particularly benefits projects requiring antibodies for low-abundance targets or applications demanding minimal background interference, such as immunohistochemistry or live-cell imaging. Clients frequently report 3-5x sensitivity improvements compared to standard approaches.

Q: What information do you need from us to start a fatty acylation antibody discovery project?

A: Essential details include: 1) Modification type and exact linkage chemistry (e.g., N-terminal myristate vs. thioester-linked palmitate), 2) Full-length or partial protein sequence containing modification sites, 3) Intended experimental applications (e.g., ELISA, flow cytometry), 4) Preferred antibody formats (species, clonality). Supplementary data, like structural models or biological samples further enhance project design.

Q: What quality control measures do you have in place throughout the antibody discovery process?

A: Our QC framework combines technical and biological validation checkpoints. Batch testing includes mass spectrometry verification for hapten integrity, SPR-based affinity measurements, and cross-reactivity profiling against 15+ common PTMs. For monoclonal lines, we perform clonality confirmation via restriction fragment analysis. All deliverables include raw screening data and performance metrics across standardized assay conditions.

Reference

1. Kim, Chung S., and Ivan A. Ross. "Regulatory Role of Free Fatty Acids (FFAs)—Palmitoylation and Myristoylation." Food and Nutrition Sciences (2013). Distributed under Open Access license CC BY 4.0, without modification.

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