Recombinant Pongo pygmaeus NADH dehydrogenase [ubiquinone] 1 beta subcomplex subunit 11, mitochondrial (NDUFB11)

What is the structural and functional role of NDUFB11 in mitochondrial complex I?

NDUFB11 serves as one of the 30 supernumerary subunits of NADH:ubiquinone oxidoreductase (complex I) of the mitochondrial respiratory chain. Structurally, it contributes to the stability of complex I assembly while functionally participating in electron transfer processes within the respiratory chain . The protein is particularly crucial for maintaining energy metabolism within mitochondria.

To study NDUFB11's structural role, researchers typically employ techniques including:

• Blue native polyacrylamide gel electrophoresis (BN-PAGE) to visualize intact respiratory complexes

• Immunoprecipitation with complex I-specific antibodies

• Cryo-electron microscopy for structural determination of the protein within the larger complex

Functional assessment requires methods such as:

• Oxygen consumption rate measurements

• NADH:ubiquinone oxidoreductase activity assays

• Mitochondrial membrane potential analysis using fluorescent probes

How does NDUFB11 gene expression differ across tissues and disease states?

Research indicates significant variation in NDUFB11 expression patterns, particularly in pathological conditions. Gene expression heatmap analysis has revealed that NDUFB11 is underexpressed in atherosclerosis accompanied by chronic stress compared to normal tissues . This differential expression pattern serves as a potential biomarker for disease progression.

Methodologically, researchers can analyze NDUFB11 expression through:

• RNA sequencing (RNA-seq) for transcriptome-wide expression analysis

• Quantitative PCR (qPCR) for targeted gene expression assessment

• Western blotting for protein-level quantification

• Immunohistochemistry for tissue-specific localization studies

What experimental approaches are recommended for studying NDUFB11 function?

When investigating NDUFB11 function, researchers should consider multiple complementary approaches:

• Gene knockdown/knockout studies: shRNA-mediated NDUFB11 knockdown in cell lines (e.g., HeLa cells) has demonstrated its essential role in complex I assembly and activity as well as cell growth and survival .

• Overexpression models: Transfection with NDUFB11-expressing vectors can help determine gain-of-function effects.

• Mitochondrial function assays:

  • Oxygen consumption rate measurements

  • ATP production quantification

  • Reactive oxygen species (ROS) detection

  • Mitochondrial membrane potential assessment

• Protein-protein interaction studies:

  • Co-immunoprecipitation

  • Proximity ligation assays

  • Yeast two-hybrid screening

These techniques should be employed in combination to obtain comprehensive insights into NDUFB11's functional significance.

How do mutations in NDUFB11 contribute to disease pathogenesis in MLS syndrome and histiocytoid cardiomyopathy?

NDUFB11 mutations have been identified in patients with Microphthalmia with linear skin defects (MLS) syndrome and histiocytoid cardiomyopathy, suggesting complex pathogenic mechanisms. The specific mutation c.262C>T (p.Arg88*) has been documented in both conditions, indicating potential shared molecular pathways .

For investigating the pathogenic mechanisms:

• Patient-derived cell models:

  • Fibroblasts or induced pluripotent stem cells (iPSCs) from patients carrying NDUFB11 mutations

  • Differentiation into disease-relevant cell types (cardiomyocytes, neural cells)

• Complex I functional assessment:

  • Enzymatic activity measurements

  • Supercomplex formation analysis

  • Electron transport chain efficiency

• Molecular consequences analysis:

  • Protein truncation effects on complex assembly

  • Altered mitochondrial ultrastructure via electron microscopy

  • Tissue-specific effects via conditional knockout animal models

The shared mutation between different phenotypic presentations suggests context-dependent effects that may be influenced by genetic background, environmental factors, or tissue-specific requirements for mitochondrial function .

What are the optimal strategies for recombinant NDUFB11 purification and functional reconstitution?

Purification and functional reconstitution of recombinant NDUFB11 present significant technical challenges due to its mitochondrial membrane localization. A comprehensive approach includes:

• Expression system optimization:

  • E. coli systems for basic structural studies

  • Mammalian or insect cell expression for post-translational modifications

  • Cell-free systems for rapid screening

• Purification protocol:

  • Detergent selection critical for membrane protein extraction

  • Affinity chromatography using tags that minimally impact function

  • Size exclusion chromatography for final purification

• Functional reconstitution:

  • Liposome incorporation with appropriate lipid composition

  • Nanodiscs for single-molecule studies

  • Co-reconstitution with other complex I components

• Quality control assessments:

  • Circular dichroism for secondary structure verification

  • Mass spectrometry for post-translational modification analysis

  • Activity assays to confirm functionality

When working with purified recombinant Pongo pygmaeus NDUFB11, researchers should avoid repeated freeze-thaw cycles and store working aliquots at 4°C for up to one week to maintain protein integrity .

How does NDUFB11 interact with other mitochondrial respiratory chain components in health and disease?

NDUFB11 functions within the intricate network of mitochondrial respiratory chain components. Protein-protein interaction (PPI) network analysis has revealed that NDUFB11 interacts with multiple components of complex I and potentially other respiratory chain complexes .

To comprehensively map these interactions:

• Physical interaction mapping:

  • Cross-linking mass spectrometry to capture dynamic interactions

  • Hydrogen-deuterium exchange mass spectrometry for interface identification

  • Cryo-electron microscopy for structural visualization

• Functional interaction assessment:

  • Respiratory supercomplex analysis under different stress conditions

  • Compensatory expression patterns in response to NDUFB11 deficiency

  • Genetic interaction screens to identify synthetic lethal partners

• Disease-relevant interaction changes:

  • Comparison of interactomes between normal and disease states

  • Identification of altered interactions in patient samples

  • Therapeutic targeting of key interaction nodes

A PPI network analysis constructed using STRING database and Cytoscape software identified 14 core genes (including NDUFB11 and NDUFS3) that interact within the mitochondrial respiratory chain, highlighting the complex interactome critical for proper function .

What are the molecular mechanisms underlying the association between NDUFB11 downregulation and atherosclerosis with chronic stress?

Recent studies have established a significant correlation between NDUFB11 underexpression and atherosclerosis complicated by chronic stress. The molecular mechanisms potentially involve:

• Mitochondrial dysfunction cascade:

  • Reduced complex I activity leading to decreased ATP production

  • Increased reactive oxygen species generation

  • Mitochondrial membrane potential collapse

  • Activation of mitochondria-dependent apoptotic pathways

• Metabolic reprogramming:

  • Shift from oxidative phosphorylation to glycolysis

  • Altered fatty acid metabolism

  • Disruption of pentose phosphate pathway

  • Impaired fructose and mannose metabolism

• Tissue-specific consequences:

  • Vascular endothelial dysfunction

  • Enhanced inflammatory response in vessel walls

  • Impaired stress response in adrenal tissues

  • Metabolic dysregulation contributing to atherosclerotic plaque formation

Experimental approaches should integrate multi-omics analysis (transcriptomics, proteomics, metabolomics) with functional studies in relevant cell types and animal models. Gene expression analysis has shown that both NDUFB11 and NDUFS3 are downregulated in atherosclerosis with chronic stress, with worse prognosis correlating with lower expression levels .

How can NDUFB11 expression patterns be leveraged for biomarker development in cardiovascular diseases?

NDUFB11 expression patterns show potential as biomarkers for cardiovascular conditions, particularly atherosclerosis complicated by chronic stress. Research indicates consistently lower expression of NDUFB11 in these conditions compared to normal controls .

To develop clinically viable biomarkers:

• Expression analysis standardization:

  • Establish reference ranges across different tissue types

  • Develop quantitative assays suitable for clinical laboratory use

  • Validate findings across diverse patient populations

• Correlation with disease progression:

  • Longitudinal studies tracking NDUFB11 expression over disease course

  • Association analysis with established clinical parameters

  • Prognostic value assessment for major adverse cardiovascular events

• Non-invasive detection methods:

  • Circulating cell-free mitochondrial DNA analysis

  • Exosome-associated NDUFB11 mRNA quantification

  • Metabolite signatures associated with NDUFB11 dysfunction

• Multimarker panel development:

  • Integration with other mitochondrial markers (including NDUFS3)

  • Algorithm development for improved diagnostic accuracy

  • Risk stratification models incorporating genetic and expression data

The Comparative Toxicogenomics Database (CTD) analysis has revealed that NDUFB11, along with NDUFS3, is associated with multiple pathological conditions including necrosis, hyperplasia, inflammation, renal disease, weight loss, memory impairment, and cognitive impairment .

What therapeutic strategies might target NDUFB11 dysfunction in mitochondrial diseases?

Given the critical role of NDUFB11 in mitochondrial function, several therapeutic approaches could address dysfunction:

• Gene therapy approaches:

  • AAV-mediated gene delivery for NDUFB11 reconstitution

  • CRISPR-based gene editing for mutation correction

  • Antisense oligonucleotides for splice-modulating therapy in specific mutations

• Small molecule interventions:

  • Complex I activity modulators

  • Mitochondrial biogenesis enhancers

  • Antioxidants targeting mitochondrial ROS

  • Metabolic bypass strategies

• Mitochondrial transplantation:

  • Direct delivery of healthy mitochondria to affected tissues

  • Induced transfer via extracellular vesicles

  • Cell-based therapies using cells with robust mitochondrial function

• Precision approaches based on mutation type:

  • Read-through therapies for nonsense mutations like c.262C>T (p.Arg88*)

  • Protein stabilization for missense mutations

  • Alternative splicing modulators for splice-site mutations

Therapeutic development should consider the tissue-specific requirements for NDUFB11 function and the differential effects of NDUFB11 deficiency across tissues, which may explain the diverse clinical presentations from similar mutations .