Discovery and structural characterization of newly identified mutations in the HBx gene of chronic HBV patients

HBV infection remains a major cause of chronic liver disease and hepatocellular carcinoma worldwide. HBx is a key regulatory protein involved in viral persistence and host-virus interactions, yet its sequence variability in Middle Eastern populations remains insufficiently characterized. In this study, the HBx region was screened in 50 Iranian patients with chronic HBV infection, and four representative samples with distinct HRM profiles were selected for sequencing. Several newly identified substitutions were detected, including two glycine substitutions (G37A and G73V) and one proline substitution (P32H), while G43D corresponded to a previously reported variant. Among these, Gly73 variants were recurrent across multiple samples. Qualitative structural modeling of full-length HBx suggested that these substitutions may contribute to subtle local conformational changes near regulatory regions; however, these observations remain preliminary and require experimental validation. Clinical and biochemical parameters showed heterogeneous patterns across cases, with Gly73 variants detected in patients with higher viral loads, although no statistically significant associations were observed due to the small sample size. All sequences belonged to genotype D. Overall, this exploratory analysis expands the existing catalog of HBx variability in a Middle Eastern cohort and highlights several newly identified substitutions that warrant further investigation in larger functional studies.