BOLA1 Human
Description
BOLA1 is fused to a 23 amino acid His-tag at N-terminus & purified by proprietary chromatographic techniques.
Product Specs
Q&A
What is BOLA1 and where is it localized in human cells?
BOLA1 is a member of the BolA protein family, which is evolutionarily widespread among eukaryotes and bacteria. In humans, BOLA1 is specifically localized to mitochondria where it functions as a regulator of thiol redox potential. The protein's bacterial homolog (BolA in Escherichia coli) causes spherical cell shape and is overexpressed during oxidative stress conditions. BOLA1 plays a significant role in maintaining normal mitochondrial morphology, particularly under oxidative stress conditions .
What is the evolutionary distribution of BOLA1 orthologs?
BOLA1 orthologs are exclusively found in aerobic eukaryotes, strongly supporting its role in controlling mitochondrial thiol redox potential. This evolutionary distribution pattern suggests BOLA1 evolved as an adaptation to aerobic metabolism and its associated challenges of oxygen toxicity and reactive oxygen species management. The absence of BOLA1 orthologs in anaerobic organisms provides important evolutionary context for understanding its specialized function in mitochondrial redox regulation .
What are the key protein interactions of BOLA1?
Studies have demonstrated that BOLA1 specifically interacts with mitochondrial monothiol glutaredoxin GLRX5. This interaction has been experimentally verified through co-purification experiments where BOLA1 (but not BOLA3) co-purified with tagged GLRX5. In yeast models, the BOLA1 homolog (Bol1) forms dimeric complexes with both monothiol glutaredoxin Grx5 and Nfu1, which influences the stability of shared Fe/S clusters .
How does BOLA1 relate to other members of the BOLA family?
In humans, BOLA1 is one of three members of the BOLA family, with BOLA3 being another mitochondrial member. While both BOLA1 and BOLA3 localize to mitochondria and are involved in Fe/S protein assembly, they exhibit distinct functions and interaction profiles. Their yeast homologs (Bol1 and Bol3) perform largely overlapping functions in iron-sulfur cluster insertion into specific mitochondrial proteins such as lipoate synthase and succinate dehydrogenase .
What cellular functions is BOLA1 involved in?
BOLA1 is primarily involved in:
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Regulating mitochondrial thiol redox potential
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Maintaining normal mitochondrial morphology under oxidative stress
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Counterbalancing the effects of glutathione depletion
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Contributing to iron-sulfur (Fe/S) protein assembly in mitochondria
When BOLA1 is knocked down, there is increased oxidation of mitochondrial thiol groups, indicating its role in maintaining the reduced state of these groups .
What methodologies are used to study BOLA1's role in mitochondrial redox regulation?
Several complementary methodologies have been employed to investigate BOLA1's role:
| Methodology | Application | Key Measures |
|---|---|---|
| Genetic manipulation | siRNA knockdown & BOLA1-GFP overexpression | Expression levels |
| Oxidative stress induction | BSO (glutathione depletion) & SNOC treatment | Redox status |
| Redox potential measurement | Thiol group oxidation assessment | Mitochondrial redox state |
| Morphology analysis | TMRM fluorescent dye visualization | Area, aspect ratio, form factor |
| Protein interaction studies | Co-purification experiments | Binding partners |
These approaches allow researchers to systematically analyze how BOLA1 affects mitochondrial function under various conditions .
How does BOLA1 affect mitochondrial morphology under oxidative stress conditions?
This protective effect is specific to BOLA1, as overexpression of BOLA3 does not prevent BSO-induced changes in mitochondrial morphology. Importantly, BSO treatment did not alter the number of mitochondria per cell, suggesting the observed morphological changes represent mitochondrial shrinkage rather than fragmentation .
What is the mechanism behind BOLA1's protective effect against glutathione depletion?
The mechanism appears to involve BOLA1's interaction with GLRX5. Although GLRX5 can reduce GSH mixed disulfides, it does so at a rate 100 times lower than dithiol glutaredoxin GLRX2 and is inefficiently reduced by GSH.
BOLA1 may enhance GLRX5's reducing activity, enabling it to:
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More efficiently reduce GSH mixed disulfides
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Be reduced more effectively by GSH itself
Through this enhanced activity, the BOLA1/GLRX5 complex likely maintains target proteins involved in mitochondrial morphology in their reduced state. This hypothesis is supported by the observation that DTT (a reducing agent) mimics BOLA1's protective effect .
How do BOLA1 and BOLA3 functionally differ in Fe/S protein biogenesis?
| Characteristic | BOLA1 | BOLA3 |
|---|---|---|
| GLRX5 interaction | Co-purifies with GLRX5 | Does not co-purify with GLRX5 |
| Effect on mitochondrial morphology | Prevents BSO-induced changes | No effect on BSO-induced changes |
| Target Fe/S proteins | Specific subset (in yeast) | Specific subset (in yeast) |
| Clinical significance | Unknown | Mutations cause MMDS2 |
In yeast, the homologs Bol1 and Bol3 have overlapping functions in Fe/S cluster insertion, but neither can replace the ISC protein Nfu1 that also participates in Fe/S protein biogenesis .
What experimental approaches have been used to determine BOLA1's interaction with GLRX5?
The BOLA1-GLRX5 interaction has been investigated using multiple approaches:
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Co-purification: BOLA1 (but not BOLA3) co-purified with tagged GLRX5, indicating a specific interaction
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Yeast comparative studies: The BOLA1 homolog (Bol1) forms dimeric complexes with both Grx5 and Nfu1
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Functional assays: BOLA1 overexpression affects cellular responses to oxidative stress and GSH depletion
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Evolutionary analysis: Co-occurrence of BOLA1 and GLRX5 in aerobic eukaryotes supports their functional interaction
Despite these findings, researchers note "we have no direct evidence for a role of GLRX5 in BOLA1's function," suggesting the need for further mechanistic studies .
How can researchers effectively manipulate BOLA1 expression to study its function?
| Approach | Methodology | Experimental Considerations |
|---|---|---|
| BOLA1 knockdown | siRNA targeting | Include non-targeting siRNA controls |
| BOLA1 overexpression | BOLA1-GFP fusion constructs | Use GFP-only and untransfected cell controls |
| Verification | Western blotting or qPCR | Confirm expression changes before functional assays |
| Phenotypic assessment | Mitochondrial morphology analysis | Use TMRM staining for visualization |
| Redox state evaluation | Measure thiol group oxidation | Compare with DTT treatment effects |
These approaches have demonstrated that BOLA1 knockdown increases mitochondrial thiol oxidation, while overexpression prevents BSO-induced morphological changes .
How do researchers reconcile contradictory findings about BOLA1 function?
Several apparent contradictions in BOLA1 research require careful interpretation:
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Dual roles in redox regulation and Fe/S protein assembly: These functions may be interconnected, as Fe/S clusters are highly sensitive to oxidative damage. BOLA1's redox regulatory role may protect Fe/S clusters or facilitate their assembly under oxidative conditions.
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GLRX5 interaction without clear functional evidence: Although BOLA1 interacts with GLRX5, there is "no direct evidence for a role of GLRX5 in BOLA1's function." This may reflect experimental limitations or indicate that GLRX5 is one of several redundant partners.
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Species-specific differences: In yeast, Bol1 and Bol3 have largely overlapping functions, yet in human cells, BOLA1 and BOLA3 appear to have distinct roles. These differences may reflect evolutionary divergence or experimental variations .
Experimental Data Table: Key BOLA1 Research Findings
Product Science Overview
- Amino Acid Sequence: The recombinant human BOLA1 protein consists of 140 amino acids (21-137 a.a) and has a molecular mass of approximately 14 kDa .
- Tagging and Purification: BOLA1 is fused to a 23 amino acid His-tag at the N-terminus and is expressed in Escherichia coli (E. coli). The protein is then purified using conventional chromatography techniques .
- Purity: The recombinant protein is highly pure, with a purity greater than 95% as determined by SDS-PAGE .
BolA proteins, including BOLA1, have been shown to be secreted into the culture medium of transfected Cos-7 cells, although they do not colocalize with the Golgi apparatus . Interestingly, the secretion of these proteins is not inhibited by Brefeldin A (BFA) treatment, which typically disrupts protein transport from the endoplasmic reticulum to the Golgi .
