Glioma is the most common and aggressive malignant tumors in central nervous system, its morbidity and mortality are both high. Lysine acetylation could alter the expression of oncogene and anti-oncogene. Thus, this study explored the potential mechanism and aimed to find new diagnostic and therapeutic methods. The Cancer Genome Atlas Glioblastoma Multiforme, Glioma related dataset were downloaded from UCSC and CGGA database respectively. Lysine acetylation-related genes (LARGs) was acquired from published literature. Differentially expressed genes (DEGs) were analyzed between GBM and control samples. LARGs (DE-LARGs) were obtained by taking the intersection of DEGs and weighted gene co-expression network analysis (WGCNA) module genes. Subsequently, enrichment analysis and protein-protein interaction (PPI) network was processed. Then, prognosis genes were selected, risk model was constructed and verified. After that, independent prognosis factors were used to predict the survival of GBM patients. Corresponding pathways and functions were analyzed between different groups. The difference of immune environment was compared. Finally, the drug prediction and regulatory network construction was performed. Prognosis genes in tumor and normal tissue were identified using immunohistochemistry. Totally 6767 DEGs were screened out. A total of 2890 module genes were identified highly correlated with lysine acetylation score by WGCNA. A total of 313 DE-LARGs were acquired by taking the intersection of DEGs and module genes. PPI network was constructed and 215 genes were obtained. Further, risk model revealed 5 genes (CD79B, STXBP4, DDHD1, FKBP1B and TRAM2) was related with overall survival (OS) of GBM patients. Kaplan-Meier survival and receiver operating characteristic curves were proved to be highly accurate both in training and validation set. Based on nomogram, riskscore was the independent prognosis factor for patients. The immune infiltration level was highly expressed in high risk group. Four drugs (PAC.1, OSI.906, WH.4.023, BMS.536924) were identified as chemotherapeutic drugs in Glioma. The transcription factors (TFs)-mRNA regulatory network was constructed and 76 TFs were obtained using TRRUST. Finally, the expression of prognostic genes in tumor was significantly higher than that in normal tissue. New prognostic genes CD79B, STXBP4, DDHD1, FKBP1B, and TRAM2 were identified for glioma through the new perspective of lysine acetylation, suggesting their importance in the development of the disease and offering potential insights for diagnosis and treatment.