Recombinant Human Vesicle transport protein GOT1B (GOLT1B)
Description
Introduction to GOLT1B
GOLT1B, also known as vesicle transport protein GOT1B, belongs to the evolutionarily conserved GOT1 protein family. This integral membrane protein plays a crucial role in the fusion of endoplasmic reticulum (ER)-derived transport vesicles with the Golgi complex . In humans, GOLT1B is encoded by the GOLT1B gene located on chromosome 12 and is expressed across various tissue types .
The protein is recognized by several alternative names in the scientific literature:
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Germ cell tumor 2
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Golgi transport 1 homolog B
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hGOT1a
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Putative NF-kappa-B-activating protein 470
Gene aliases include CGI-141, GCT2, GOLT1B, GOT1, GOT1A, GOT1B, HDCMA39P, UNQ432/PRO793, and YMR292W, with human GOLT1B assigned the UniProt ID Q9Y3E0 and Entrez Gene ID 51026 .
Role in COPII-Mediated Vesicle Transport
GOLT1B plays a critical role in the COPII (Coat Protein Complex II)-mediated protein export from the endoplasmic reticulum. COPII represents the primary machinery responsible for anterograde transport of newly synthesized proteins from the ER to other endomembrane compartments in eukaryotes .
Research using rice (Oryza sativa) has demonstrated that GOT1B regulates COPII vesicle formation at ER exit sites (ERESs), facilitating protein trafficking. In the rice glutelin precursor accumulation4 (gpa4) mutant, which lacks functional GOT1B, researchers observed:
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Accumulation of 57-kD glutelin precursors
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Formation of two types of ER-derived abnormal structures
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Altered distribution of Sar1 in the endomembrane system
These findings suggest that GOT1B functions in the early stages of COPII vesicle formation, potentially before the budding process occurs.
Protein Interactions
GOLT1B engages in specific protein interactions that facilitate its function in vesicular transport. Key interactions include:
| Interaction Partner | Detection Method | Functional Significance |
|---|---|---|
| Sec23 (COPII component) | Yeast two-hybrid, BiFC, Co-IP | Forms prebudding complexes |
| Sar1b | Co-immunoprecipitation | Present in same complex(es) in vivo |
| TBK1 | Western blot, Dual luciferase assay | Modulates NF-κB pathway |
Studies have confirmed that GOLT1B directly interacts with Sec23, a component of the COPII coat. This interaction has been validated through multiple experimental approaches, including yeast two-hybrid, bimolecular fluorescence complementation (BiFC), and co-immunoprecipitation assays .
Additionally, GOLT1B, Sec23c, and Sar1b are present in the same complex(es) in vivo, suggesting that GOLT1B functions at the stage of prebudding complex formation rather than after COPII vesicle release .
Production and Characteristics
Recombinant human GOLT1B is produced through various expression systems for research applications. One approach involves cell-free protein synthesis (CFPS) systems, which offer advantages in producing membrane proteins like GOLT1B. Commercially available recombinant GOLT1B typically includes:
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Full-length proteins (amino acids 1-138)
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Control fragments (such as amino acids 110-135)
Production typically involves one-step affinity chromatography purification, with purity levels generally exceeding 70-80% as determined by SDS-PAGE, Western Blot, and analytical SEC (HPLC) .
The protein's concentration is measured using absorbance at 280nm against a specific reference buffer, with the absorption coefficient determined using tools such as Expasy's ProtParam .
Applications in Research
Recombinant human GOLT1B serves multiple purposes in scientific research:
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Antibody Validation: Recombinant GOLT1B control fragments are used for blocking experiments with corresponding antibodies (e.g., PA5-65988, PA5-64276). For immunohistochemistry/immunocytochemistry and Western blot experiments, a 100x molar excess of the protein fragment control is typically recommended, with pre-incubation of the antibody-protein control fragment mixture for 30 minutes at room temperature .
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Functional Studies: Recombinant GOLT1B enables investigations into protein transport mechanisms between the ER and Golgi apparatus.
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Disease Research: It facilitates studies on GOLT1B's role in various pathological conditions, particularly cancer development and progression.
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Interaction Analysis: Recombinant GOLT1B is used to validate and characterize protein-protein interactions within the vesicular transport machinery.
It is important to note that commercially available recombinant human GOLT1B is intended for research use only and not for diagnostic procedures or resale without express authorization .
Role in Cancer Progression
Recent research has implicated GOLT1B in multiple cancer types, suggesting its potential as both a prognostic biomarker and therapeutic target.
Breast Cancer
Multi-omics analyses have identified GOLT1B as a potential prognostic marker in breast cancer. The expression of GOLT1B is significantly higher in breast cancer tissues compared to normal mammary tissues. Key findings include:
Cervical Cancer
GOLT1B has been found to promote the progression of cervical cancer (cervical squamous cell carcinoma and endocervical adenocarcinoma—CESC) through specific molecular mechanisms:
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Upregulation in cervical cancer tissue compared to normal tissue
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Association with poor prognosis
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Promotion of cervical cancer viability and migration both in vitro and in vivo
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Enhancement of the NF-κB pathway through interaction with TANK-binding kinase 1 (TBK1)
Other Cancers
GOLT1B has also been implicated in:
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Colorectal cancer: Overexpression can elevate cell membrane levels of DVL2, activating the Wnt/β-catenin pathway and inducing epithelial-mesenchymal transformation; it also promotes migration and invasion via inducing T lymphocyte apoptosis
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Lung adenocarcinoma: Patients with GOLT1B amplifications show poorer prognosis
Immune Microenvironment Regulation
A particularly intriguing aspect of GOLT1B function relates to its role in regulating the tumor immune microenvironment. Studies have revealed significant correlations between GOLT1B expression and immune cell infiltration patterns in various cancers .
In breast cancer, GOLT1B expression correlates with:
| Positive Correlation | Negative Correlation |
|---|---|
| Macrophages | Monocytes |
| M0 macrophages | CD8+ T cells |
| M2 macrophages | CD4+ T cells |
| Neutrophils | Regulatory T cells |
| iTreg cells | Helper T cells |
| nTreg cells | Plasma cells |
| Dendritic cells | NK cells |
| Central memory T cells | Gamma delta T cells |
| Type 1 helper T cells | Helper follicular T cells |
Additionally, GOLT1B expression positively correlates with several immune checkpoint molecules, including CD274, TIGIT, and CTLA4 . These findings suggest that GOLT1B may influence cancer progression partly through modulating the immune microenvironment.
Current Research and Future Directions
Current research on GOLT1B focuses on elucidating its precise mechanisms in vesicular transport and disease pathogenesis. Several models have been proposed regarding its function in COPII vesicle formation.
One speculative model suggests that GOT1B, localized to the ERESs, works cooperatively with Sar1 to facilitate the recruitment of the Sec23/Sec24 heterodimer, forming prebudding complexes containing preloaded cargos. Subsequently, the Sec13/Sec31 heterotetramer is recruited to form the outer coat of the COPII vesicles before vesicle budding and fusion with the cis-Golgi apparatus .
Future research directions may include:
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Detailed structural studies of GOLT1B to elucidate its membrane topology and interaction interfaces
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Investigation of GOLT1B as a potential therapeutic target in cancer treatment
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Exploration of its role in regulating immune responses beyond the cancer microenvironment
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Development of GOLT1B-targeted therapies that could modulate vesicular transport in disease states
Product Specs
Note: We will prioritize shipping the format currently in stock. However, if you have a specific format requirement, please indicate it in your order. We will prepare the product according to your request.
Note: All our proteins are shipped with standard blue ice packs. If dry ice shipping is required, please inform us in advance, as additional fees will apply.
Generally, the shelf life of the liquid form is 6 months at -20°C/-80°C. The shelf life of the lyophilized form is 12 months at -20°C/-80°C.
Target Background
Q&A
Basic Research Questions
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What is GOLT1B and what is its primary function in cellular transport systems?
GOLT1B (Golgi Transport 1B) is an evolutionarily conserved integral membrane protein that functions in the early secretory pathway. It plays a critical role in regulating COPII-mediated protein export from the endoplasmic reticulum (ER) to other endomembrane compartments. While initially thought to be involved in the fusion of uncoated COPII vesicles to the Golgi in yeast studies, more recent evidence suggests that GOLT1B primarily functions in the assembly or budding stage of COPII vesicles by interacting with components of the COPII coat machinery . GOLT1B localizes to Golgi-associated ER exit sites (ERESs) and facilitates the anterograde transport of secretory proteins in eukaryotic cells . This protein appears to be functionally conserved across species, from yeast to plants to humans, indicating its fundamental importance in cellular transport processes.
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How does GOLT1B relate to the COPII vesicle transport system?
GOLT1B interacts directly with the COPII coat machinery, particularly with the Sec23 component. Through yeast two-hybrid (Y2H) assays, bimolecular fluorescence complementation (BiFC), and co-immunoprecipitation (co-IP) experiments, researchers have demonstrated that GOLT1B specifically interacts with Sec23 isoforms . Furthermore, co-IP assays have shown that Sar1b (another COPII component), GOT1B, and Sec23c can be present in the same complex(es) in vivo . These interactions suggest that GOLT1B functions before the budding of COPII vesicles, likely participating in the formation or stability regulation of the prebudding complex. In GOLT1B-deficient cells, the fractionation pattern of prebudding complex components (Sar1b and Sar1c) is altered, and there is a reduction in the amount of modified Sec23c protein immunoprecipitated by anti-Sar1b antibodies . This evidence collectively supports GOLT1B's role in regulating COPII vesicle formation.
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What cellular phenotypes are observed when GOLT1B function is disrupted?
Disruption of GOLT1B function leads to several observable cellular phenotypes related to protein transport. In rice mutants lacking functional GOT1B (gpa4 mutant), researchers observed the accumulation of 57-kD glutelin precursors and the formation of two types of ER-derived abnormal structures . The distribution pattern of ER exit sites (ERESs) is also significantly altered in GOLT1B-deficient cells, changing from a dispersed distribution in wild-type cells to a more concentrated pattern in mutant cells . This change likely reflects disrupted recycling of COPII vesicles between the ER and Golgi apparatus. Additionally, in GOLT1B-deficient cells, the recycling of COPII coat components appears to be affected, leading to blockage of these components in the ERESs . These phenotypes collectively indicate that GOLT1B plays an essential role in maintaining normal protein trafficking from the ER to the Golgi.
Experimental Design Questions
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How should researchers design experiments to distinguish between GOLT1B's direct effects on COPII trafficking versus secondary consequences?
Designing experiments to distinguish direct from indirect effects of GOLT1B requires careful temporal and mechanistic controls. Acute depletion systems, such as auxin-inducible degron approaches, allow researchers to observe immediate consequences of GOLT1B loss before compensatory mechanisms develop. Rescue experiments introducing wild-type or mutant GOLT1B (particularly mutations in regions mediating Sec23 interaction) into GOLT1B-deficient backgrounds can establish causality and identify essential functional domains . Structure-function analyses with chimeric proteins or domain deletions can pinpoint which GOLT1B regions are necessary for specific aspects of COPII trafficking. In vitro reconstitution assays with purified components represent a powerful approach to test whether GOLT1B directly influences COPII coat assembly rates or stability . Time-resolved microscopy tracking the sequential recruitment of COPII components can determine if GOLT1B affects early (Sar1, Sec23/24) or late (Sec13/31) stages of coat assembly. Cargo-specific trafficking assays can reveal whether GOLT1B selectively influences certain cargo classes, suggesting direct effects on cargo selection versus general trafficking defects. These approaches collectively help establish where and how GOLT1B directly interfaces with the COPII machinery.
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What considerations should researchers take into account when developing GOLT1B as a potential cancer biomarker?
Developing GOLT1B as a cancer biomarker requires addressing several key considerations. Researchers must establish clear standardization of GOLT1B detection methods, whether through immunohistochemistry, RT-PCR, or other approaches, to ensure reproducibility across laboratories . Analytical validation should include assessment of sensitivity, specificity, and reproducibility across different sample types and preservation methods. Clinical validation requires large, well-characterized patient cohorts with adequate statistical power, representing diverse populations and cancer subtypes . Since GOLT1B expression has been linked to breast cancer prognosis, researchers should determine optimal cut-off values that meaningfully stratify patients into risk groups . Context-specific validation is crucial - GOLT1B's prognostic value may vary by cancer subtype, treatment history, or when combined with other biomarkers. Researchers should assess GOLT1B's independent prognostic value through multivariate analyses accounting for established clinicopathological factors . For clinical utility, studies should evaluate whether GOLT1B-based stratification affects treatment decisions or outcomes. Finally, practical implementation factors including assay cost, complexity, and turnaround time need consideration for clinical translation. Following these considerations will help establish whether GOLT1B can serve as a robust, clinically useful biomarker.
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How can researchers investigate the evolutionary conservation of GOLT1B function across different species?
Investigating GOLT1B's evolutionary conservation requires an integrated comparative approach. Sequence analysis across species, from yeast GOT1p to plant GOT1B to mammalian GOLT1B, can identify conserved domains and motifs that may indicate functional importance . Structural predictions and comparative modeling provide insights into conservation at the three-dimensional level. Complementation studies, where human GOLT1B is expressed in plant or yeast mutants lacking functional GOT1B/GOT1p, can test functional conservation directly . Researchers can perform comparative interaction studies to determine if GOLT1B's binding partners (like Sec23) are consistent across species; evidence suggests that yeast and human versions of GOLT1B can interact with their corresponding Sec23 proteins . Comparative localization studies using fluorescently tagged GOLT1B from different species can reveal whether subcellular targeting is conserved. When examining phenotypes, researchers should compare cellular consequences of GOLT1B deficiency across species, looking for common patterns in secretory pathway disruption . This multi-faceted approach can determine which aspects of GOLT1B function represent ancient, conserved roles in the secretory pathway versus species-specific adaptations.
