Leaky expression of PRMT1 in Escherichia coli and redox regulation of its histone methyltransferase activity

Protein arginine methyltransferase 1 (PRMT1) is a major histone and non-histone arginine methyltransferase whose activity is important for gene regulation and genome stability. In this study, we investigated how expression conditions and redox environment affect recombinant human PRMT1 produced in Escherichia coli BL21(DE3). We hypothesized that (i) basal T7 promoter activity would generate measurable PRMT1 protein even without IPTG induction, and (ii) stronger reducing conditions would enhance PRMT1-mediated histone methylation in a DTT-dependent manner, whereas β-mercaptoethanol would have little effect. Small-scale induction tests using a pET28a(+) construct revealed a clear PRMT1 band in whole-cell lysates at all IPTG concentrations, including 0 mM, and Ni-NTA purification yielded comparable amounts of PRMT1 across tested conditions, confirming leaky expression. In vitro histone methyltransferase assays using recombinant histone octamer showed that H4R3 asymmetric dimethylation increased with PRMT1 amount, saturated at higher enzyme levels, and was abolished by the catalytic-dead mutant PRMT1(G98R). Titration of DTT demonstrated a dose-dependent increase in PRMT1 activity that
plateaued at higher concentrations, whereas replacing DTT with β-mercaptoethanol, even at much higher concentrations, did not further stimulate methylation. Together, these findings support our hypothesis that leaky T7 expression is sufficient for PRMT1 production in BL21(DE3) and that PRMT1 catalytic activity is strongly regulated by the strength of the DTT-dependent reducing environment.