Background and objectives <p>The clinical behavior and molecular mechanisms of hepatocellular carcinoma (HCC) are intricate and highly variable, posing challenges for identifying novel targets in clinical research. The gene eukaryotic translation initiation factors 5B (EIF5B) plays a role in synthesize translation initiation complexes during the synthesis of eukaryotic proteins.</p> Materials and methods <p>We conducted a comprehensive bioinformatics analysis using public databases including Tumor Immune Estimation Resource (TIMER), UALCAN, Kaplan-Meier plotter, LinkedOmics, and Gene Expression Profiling Interactive Analysis (GEPIA2).</p> Results <p>EIF5B expression in HCC samples was significantly higher than in normal liver tissue; additionally, EIF5B was highly in primary tumors, various stages of cancer and grades of tumor, and status of nodal metastasis. Higher EIF5B expression was also associated with diagnosis. EIF5B expression showed a positive correlation with B cells macrophages, myeloid dendritic cells, neutrophils, and CD4⁺ T cells. In the analysis of EIF5B co-expression networks, positively related genes of EIF5B were associated with a high hazard ration in different types of cancer, including HCC. In biological function of EIF5B, mainly plays including protein localization to chromosome, rRNA metabolic process, and ncRNA processing, etc.</p> Conclusion <p>These results suggest that EIF5B may serve as a novel biomarker with prognostic relevance in HCC and provide insights into tumor immunology in HCC.</p>