GARS Human

What is GARS and what are its primary functions in human cells?

GARS (Glycyl-tRNA synthetase) is an (alpha)2 dimer that belongs to the class II family of tRNA synthetases. It serves dual critical functions in human cellular processes:

• Its primary role is catalyzing the attachment of glycine to tRNA(Gly) during protein translation

• It produces diadenosine tetraphosphate (Ap4A) through direct condensation of two ATP molecules

Ap4A functions as a universal pleiotropic signaling molecule required for various cell regulation pathways. Additionally, GARS has been identified as a target of autoantibodies in human autoimmune diseases, particularly polymyositis and dermatomyositis .

What is the molecular structure and characterization of human GARS?

Human GARS protein has the following structural characteristics:

The recombinant form often used in laboratory research typically includes a His-tag at the N-terminus to facilitate purification. SDS-PAGE analysis confirms a purity greater than 85% for properly prepared samples .

How are mutations in the GARS gene linked to neuropathic disorders?

Mutations in the GARS gene have been identified as causative factors in Charcot-Marie-Tooth neuropathy type 2D (CMT2D), an axonal form of inherited peripheral neuropathy. In a study of Japanese patients with axonal CMT, researchers identified a novel heterozygous Pro244Leu (c.893C>T) mutation in a patient showing adolescent onset and early upper limb involvement .

The prevalence of GARS mutations appears to vary by population. For instance, in the Japanese cohort study involving 89 patients with axonal CMT, only one patient carried a GARS mutation, suggesting that while GARS mutations can cause CMT2, they represent a relatively rare genetic cause in this population .

How does GARS relate to the Genetic Addiction Risk Score in neurogenetics?

It's important to distinguish between GARS (Glycyl-tRNA synthetase) and GARS™ (Genetic Addiction Risk Score), which represent entirely different scientific concepts despite sharing the same acronym:

• GARS (Glycyl-tRNA synthetase): The aminoacyl-tRNA synthetase enzyme discussed throughout this document

• GARS™ (Genetic Addiction Risk Score): A panel of genes and their polymorphisms used to assess genetic predisposition to addiction and Reward Deficiency Syndrome (RDS)

The Genetic Addiction Risk Score examines genes related to dopaminergic function, including DRD2, DRD3, DRD4, MOA-A, COMT, DAT1, 5HTTLLR, OPRM1, and GABRA3. This approach aims to identify individuals with a hypodopaminergic trait that may predispose them to substance use disorders and other reward-seeking behaviors .

Research on the Genetic Addiction Risk Score has demonstrated promising applications in addiction medicine. In one study, researchers were able to blindly describe lifetime Reward Deficiency Syndrome behaviors in a recovering addict (17 years sober) solely by analyzing the individual's GARS™ data . This suggests potential utility in personalized addiction treatment approaches.

What are the optimal protocols for producing recombinant human GARS protein?

The production of high-quality recombinant human GARS protein for research purposes typically follows this methodological approach:

• Expression System: E. coli is the preferred expression system for human GARS recombinant protein production

• Construct Design: The protein construct usually contains amino acids 43-289 of the native protein fused to a 23-amino acid His-tag at the N-terminus

• Purification Method: Proprietary chromatographic techniques utilizing the His-tag for affinity chromatography

• Quality Control:

  • Molecular mass confirmation by MALDI-TOF (expected: 30 kDa)

  • Purity assessment by SDS-PAGE (typically >85%)

  • Concentration determination by Bradford assay (typically 0.5 mg/ml)

The purified protein is generally formulated in 20mM Tris-HCl buffer (pH 8.0) containing 0.15M NaCl, 10% glycerol, and 1mM DTT to maintain stability and functionality .

What are the recommended storage conditions for maintaining GARS protein stability?

For optimal retention of GARS protein activity and structural integrity, the following storage protocols are recommended:

These conditions have been experimentally validated to preserve protein functionality and structural integrity for research applications .

How can functional assays be designed to evaluate the impact of GARS mutations?

Although the search results don't provide specific protocols, a comprehensive approach to assessing GARS mutations would typically include:

• Aminoacylation Activity Assays: Measuring the enzyme's capacity to attach glycine to tRNA(Gly)

• Diadenosine Tetraphosphate Synthesis: Evaluating Ap4A production through direct ATP condensation

• Protein Folding and Stability Assessments: Using thermal shift assays or circular dichroism

• Protein-Protein Interaction Studies: Identifying altered binding with cellular partners

• Subcellular Localization: Examining changes in cellular distribution of mutant GARS

These functional assays provide critical insights into how specific mutations affect GARS activity and may contribute to pathological conditions like Charcot-Marie-Tooth neuropathy.

What approaches are most effective for studying GARS autoantibodies in autoimmune disorders?

GARS has been identified as a target of autoantibodies in human autoimmune diseases, particularly polymyositis and dermatomyositis . Research methods for investigating these autoantibodies would typically include:

• Immunoprecipitation Assays: To detect and quantify autoantibodies against GARS

• ELISA Development: For high-throughput screening and quantification

• Epitope Mapping: To identify the specific regions of GARS targeted by autoantibodies

• Functional Inhibition Studies: To determine how autoantibodies affect GARS enzymatic activity

• Animal Models: To investigate the pathogenic role of anti-GARS autoantibodies in vivo

Understanding the relationship between GARS autoantibodies and disease manifestation requires a multifaceted experimental approach combining these techniques.

What genetic screening approaches are recommended for identifying GARS mutations in patients?

Based on research practices in the field, comprehensive genetic screening for GARS mutations in CMT patients should follow this methodological workflow:

• Patient Selection: Focus on individuals with clinical presentations consistent with CMT2D, particularly those with adolescent onset and early upper limb involvement

• DNA Extraction: From peripheral blood using standard protocols

• PCR Amplification: Of all GARS exons and exon-intron boundaries

• Sequencing Method: Direct sequencing of PCR products or next-generation sequencing approaches

• Variant Analysis: Comparison with reference sequences and population databases

• Functional Validation: Of novel variants using in vitro studies to confirm pathogenicity

This systematic approach was effective in identifying the novel Pro244Leu (c.893C>T) mutation in a Japanese patient with axonal CMT .

How might therapeutic strategies for GARS-related disorders be developed?

While the search results don't specifically address therapeutic approaches for GARS-related disorders, potential strategies based on current understanding would include:

• Gene Therapy: Delivery of wild-type GARS to affected tissues

• RNA-Based Therapies: Antisense oligonucleotides to modulate splicing or expression

• Small Molecule Screening: To identify compounds that enhance residual GARS activity

• Protein Replacement: Administration of functional recombinant GARS

• Cellular Pathway Modulation: Targeting downstream effects of GARS dysfunction

Development of these therapeutic approaches would require comprehensive understanding of the molecular mechanisms underlying GARS-related pathologies.

What are the current gaps in understanding GARS function in human disease?

Despite significant advances in GARS research, several critical knowledge gaps remain that warrant further investigation:

• The complete spectrum of GARS mutations and their prevalence across different populations

• Detailed molecular mechanisms by which GARS mutations lead to neurodegeneration

• The role of GARS in non-canonical cellular functions beyond aminoacylation

• Interactions between GARS and other cellular components in health and disease

• Environmental factors that may influence GARS expression and function

Addressing these knowledge gaps will require interdisciplinary approaches combining genetics, biochemistry, cell biology, and clinical research.

How might emerging technologies advance GARS research in the next decade?

Emerging technologies that could substantially advance GARS research include:

• CRISPR-Cas9 Gene Editing: For creating precise disease models and potential therapeutic applications

• Single-Cell Technologies: To understand cell-specific impacts of GARS dysfunction

• Cryo-EM Structural Analysis: For high-resolution visualization of GARS-tRNA interactions

• Proteomics Approaches: To map the complete GARS interactome

• Patient-Derived iPSCs: For personalized disease modeling and drug screening

These technologies will enable more precise investigation of GARS function and dysfunction, potentially leading to novel therapeutic approaches for GARS-related disorders.