The PCCB Knockout Hep-G2 Cell Line is a genetically engineered human liver model in which the PCCB gene has been disrupted by CRISPR/Cas9-mediated genome editing. This knockout cell line, derived from the Hep-G2 host, provides a powerful in vitro system for investigating the consequences of propionyl-CoA carboxylase deficiency. By eliminating functional PCCB expression, the model enables researchers to dissect propionate catabolism and its integration with mitochondrial energy metabolism, offering a physiologically relevant platform for metabolic disease studies.
Hep-G2 cells are a well-characterized human hepatocellular carcinoma line that retains many liver-specific functions, including epithelial morphology, adherent growth, and expression of hepatocyte markers. These cells are widely employed as a model for hepatic drug metabolism, lipid processing, and xenobiotic responses. Their hepatocyte-like features make them particularly suitable for studying inherited metabolic disorders with major hepatic involvement, such as propionic acidemia, within a controlled cellular environment.
PCCB encodes the beta subunit of propionyl-CoA carboxylase (PCCase), a biotin-dependent mitochondrial enzyme that catalyzes the ATP-dependent carboxylation of propionyl-CoA to D-methylmalonyl-CoA. The functional carboxylase is a multimeric complex in which the beta subunit interacts with the alpha subunit (PCCA) and the biotin carboxyl carrier protein. PCCB activity is transcriptionally regulated by PPARA and responsive to biotin availability and mitochondrial stress signals. Downstream, the product methylmalonyl-CoA is isomerized to succinyl-CoA by methylmalonyl-CoA mutase, an adenosylcobalamin-dependent reaction, thereby feeding into the TCA cycle and gluconeogenesis. Loss of PCCB disrupts this pathway, leading to accumulation of propionyl-CoA and toxic metabolites, impaired anaplerosis, and secondary mitochondrial dysfunction.
In the Hep-G2 context, PCCB knockout recapitulates key metabolic derangements of propionic acidemia at the hepatocellular level. The absence of functional PCCB impairs the clearance of propionyl-CoA derived from branched-chain amino acid oxidation, odd-chain fatty acid degradation, and gut fermentation, causing substrate overload and energy deficit. This cell line enables detailed examination of how defective propionate metabolism alters TCA cycle flux, NADH/NAD+ ratios, and gluconeogenic capacity, and how these changes contribute to acidosis, hyperammonemia, and organelle stress characteristic of the human disease.
This model is ideally suited for a wide range of research applications, including mechanistic studies of organic acidurias, drug screening for metabolic acidosis, and validation of gene therapy strategies targeting hepatic metabolism. Researchers can characterize PCCB disruption by Western blotting and RT-qPCR, assess residual carboxylase activity via enzyme assays, profile metabolic intermediates through metabolomics, and evaluate mitochondrial respiration under various substrate conditions. Combining this knockout line with biotin supplementation or candidate therapeutics allows dissection of compensatory pathways and assessment of rescue efficacy. For further details or to order the PCCB Knockout Hep-G2 Cell Line, please contact Ascent Research.
