NDUFS5 Human

What is the basic structure and size of human NDUFS5?

Human NDUFS5 (NADH:Ubiquinone Oxidoreductase Subunit S5) is encoded by an open reading frame of 321 base pairs that translates into a 106 amino acid protein with a calculated molecular mass of approximately 12.5 kDa . The protein belongs to the NADH dehydrogenase (ubiquinone) iron-sulfur protein family . As a subunit of Complex I, NDUFS5 is located within the inner mitochondrial membrane, primarily functioning as an accessory subunit rather than a catalytic component .

What is the primary function of NDUFS5 in cellular metabolism?

NDUFS5 serves as an accessory subunit of the mitochondrial membrane respiratory chain NADH dehydrogenase (Complex I) . While not directly involved in catalysis, it plays an essential role in Complex I assembly and stability, facilitating the transfer of electrons from NADH to the respiratory chain . The immediate electron acceptor for the enzyme is believed to be ubiquinone. NDUFS5 contributes to critical cellular pathways including respiratory electron transport, ATP synthesis by chemiosmotic coupling, and Complex I biogenesis .

Where is NDUFS5 localized within mitochondria?

Based on structural and comparative studies, NDUFS5 is localized to the mitochondrial inner membrane and intermembrane space . It shows a characteristic cytoplasmic expression pattern that appears granular when visualized through immunohistochemistry techniques . This specific localization is crucial for its role in Complex I assembly and electron transport chain function.

What is the tissue expression pattern of NDUFS5 in humans?

NDUFS5 mRNA is expressed ubiquitously across human tissues, with relatively higher expression observed in heart, skeletal muscle, liver, kidney, and fetal heart . In brain tissue, it shows general cytoplasmic expression with a granular pattern characteristic of mitochondrial proteins . The ubiquitous expression pattern correlates with its fundamental role in cellular energy metabolism, with enrichment in tissues having high energy demands .

How is NDUFS5 expression regulated during development and in different physiological states?

While comprehensive developmental regulation data is limited in the provided search results, research indicates that NDUFS5 shows notable expression in fetal heart tissue . This suggests potential developmental regulation that aligns with the increasing energy demands during organogenesis. For experimental investigation of NDUFS5 regulation, researchers should consider using quantitative PCR analysis of tissues at different developmental stages, combined with immunohistochemistry to visualize expression patterns spatiotemporally.

What methodologies are most effective for measuring NDUFS5 expression levels?

For quantitative expression analysis of NDUFS5, researchers should employ RT-qPCR with validated primers specific to NDUFS5 transcripts. Western blotting using antibodies against NDUFS5 (such as those identified by UniProtKB/Swiss-Prot: O43920) can confirm protein expression levels. For spatial expression patterns, immunohistochemistry with anti-NDUFS5 antibodies provides insights into cellular and subcellular localization . RNA-seq data from resources like the Human Protein Atlas can provide comparative expression profiles across multiple tissues and cell types.

What diseases are associated with NDUFS5 mutations or dysfunction?

Diseases associated with NDUFS5 include Cardiomyopathy, Familial Hypertrophic, 2 and Mitochondrial DNA Depletion Syndrome 9 . Complex I deficiency disorders may also involve NDUFS5, though early mutation screening (1999) in twenty isolated enzymatic complex I-deficient patients revealed no mutations or polymorphisms in this gene . More recent research may have identified additional disease associations not covered in the provided search results.

What are the mechanisms by which NDUFS5 mutations lead to mitochondrial dysfunction?

NDUFS5 mutations potentially impact mitochondrial function through several mechanisms: disrupting Complex I assembly, reducing stability of the complex, impairing electron transport efficiency, or affecting interactions with other respiratory chain components . As an accessory subunit, mutations in NDUFS5 may not directly affect catalytic activity but instead compromise the structural integrity of Complex I. The specific position of mutations can determine whether they affect protein-protein interactions, protein stability, or proper localization within the mitochondrial membrane .

What experimental approaches can detect pathogenic NDUFS5 variants in clinical samples?

For detecting pathogenic NDUFS5 variants, researchers should implement a multi-layered approach: (1) DNA sequencing (preferably next-generation sequencing) targeting the NDUFS5 gene located on chromosome 1 ; (2) Analysis of Complex I assembly and activity in patient samples using blue native polyacrylamide gel electrophoresis (BN-PAGE) to assess complex integrity; (3) Functional studies in cellular models to determine the impact of identified variants on mitochondrial respiration, reactive oxygen species production, and ATP synthesis; (4) Structural analysis to predict how mutations might affect interactions with other Complex I subunits, particularly NDUFS1 which has close contacts with NDUFS5 .

What are the most effective methods for studying NDUFS5's role in Complex I assembly?

To investigate NDUFS5's role in Complex I assembly, researchers should employ: (1) CRISPR/Cas9-mediated knockout or knockdown of NDUFS5 followed by analysis of Complex I formation using BN-PAGE; (2) Proximity labeling techniques such as BioID or APEX to identify direct interaction partners during assembly; (3) Pulse-chase experiments with radiolabeled amino acids to track the temporal sequence of Complex I assembly; (4) Cryo-electron microscopy to visualize structural changes in Complex I when NDUFS5 is absent or mutated. These approaches provide complementary insights into both the kinetics and structural aspects of NDUFS5's contribution to Complex I biogenesis.

How can researchers effectively model NDUFS5 mutations to study pathophysiology?

For modeling NDUFS5 mutations, researchers should consider: (1) Patient-derived fibroblasts or induced pluripotent stem cells (iPSCs) differentiated into relevant cell types (cardiomyocytes, neurons); (2) CRISPR/Cas9-engineered cell lines with specific NDUFS5 mutations; (3) Transgenic animal models (mice, zebrafish) with corresponding mutations; (4) In vitro reconstitution of Complex I with mutant NDUFS5 proteins. These models enable investigation of how mutations affect protein stability, Complex I assembly, mitochondrial function, and cellular pathophysiology. Structural analysis using the human Complex I structure (Protein Data Bank file 5xtd) can help predict how specific mutations might disrupt interactions with other subunits .

What are the challenges in studying NDUFS5 interactions within the mitochondrial membrane?

Studying NDUFS5 interactions presents several methodological challenges: (1) The hydrophobic environment of the mitochondrial membrane complicates protein extraction while maintaining native interactions; (2) The complex architecture of respiratory complexes requires specialized techniques for preserving structural integrity; (3) Distinguishing direct from indirect interactions within densely packed supercomplexes. Researchers should employ crosslinking mass spectrometry (XL-MS), hydrogen-deuterium exchange mass spectrometry (HDX-MS), or cryo-electron microscopy for structural studies. For interaction mapping, proximity labeling combined with mass spectrometry can identify neighboring proteins in their native environment.

How conserved is NDUFS5 across species, and what does this reveal about its function?

NDUFS5 shows significant evolutionary conservation, with 81.0% identity between human and bovine NDUFS5 at the cDNA level and 74.5% identity at the amino acid level . The zebrafish ortholog shares functional predictions regarding mitochondrial respiratory chain complex I assembly and localization . This high degree of conservation suggests essential functions that have been maintained throughout vertebrate evolution. Comparative analysis of NDUFS5 across species can reveal conserved domains critical for function versus regions that may have undergone adaptive evolution.

What can cross-species comparisons tell us about NDUFS5 structure-function relationships?

Cross-species comparative analysis can identify: (1) Conserved residues likely essential for core functions; (2) Species-specific variations that may reflect adaptations to different metabolic demands; (3) Structural motifs necessary for proper Complex I assembly across diverse organisms. The zebrafish ndufs5 shows predicted involvement in mitochondrial respiratory chain complex I assembly similar to its human counterpart , suggesting functional conservation despite some sequence divergence. Researchers can leverage these comparisons to identify critical functional domains and predict the impact of specific mutations.

How can model organisms advance our understanding of NDUFS5 function?

Model organisms offer valuable systems for studying NDUFS5 function: (1) Zebrafish models provide opportunities for in vivo visualization of mitochondrial dynamics and developmental effects of ndufs5 mutations ; (2) Mouse models enable comprehensive physiological assessment of tissue-specific consequences; (3) Cell culture models from various species allow comparative biochemical analyses. When designing experiments with model organisms, researchers should consider species-specific differences in mitochondrial biology and compensatory mechanisms that might not be present in humans.

What is known about NDUFS5's protein-protein interactions within Complex I?

NDUFS5 interacts with several Complex I subunits, with particularly important interactions with NDUFS1 . Specifically, Lys45 of NDUFS5 is found at the interface with NDUFS1 and has close contacts with several NDUFS1 residues, including Gly376, Asp380, and Ser672 . These interactions are critical for the structural stability of Complex I. Disruption of these interactions, such as through mutations, can destabilize the binding of NDUFS5 to NDUFS1 and potentially compromise Complex I integrity.

How does NDUFS5 contribute to the assembly and stability of Complex I?

As an accessory subunit, NDUFS5 contributes to Complex I assembly and stability rather than catalytic activity . Its strategic location in the mitochondrial inner membrane and intermembrane space positions it to facilitate proper assembly of Complex I components . While specific assembly steps involving NDUFS5 are not fully elucidated in the provided search results, disruption of NDUFS5 can potentially affect the expression, import, or stability of other subunits in the complex . Researchers investigating these processes should consider pulse-chase experiments and analysis of assembly intermediates to determine the precise temporal position of NDUFS5
incorporation during Complex I biogenesis.

What methodological approaches can resolve contradictory findings about NDUFS5 function?

To address contradictory findings about NDUFS5 function, researchers should implement: (1) Multiple complementary techniques (biochemical, structural, genetic) to cross-validate observations; (2) Carefully controlled knockout/knockdown studies with rescue experiments to confirm specificity; (3) Time-resolved analyses to distinguish primary from secondary effects; (4) Tissue-specific and context-dependent investigations that account for potential compensatory mechanisms. When contradictory results emerge, systematic investigation of cell type differences, experimental conditions, and methodological variations can often reconcile apparent discrepancies and lead to more nuanced understanding of NDUFS5 function.