This study investigates the impact of G-quadruplex (G4) structures in the 5' untranslated region (5'UTR) of bacterial mRNA on translation efficiency. Using an in vitro translation system and E. coli, the researchers found that G4 structures in the 5'UTR enhance translation in a size-dependent manner. Longer loops within G4 structures and the presence of a hairpin upstream of the G4 further increase translation efficiency. The effect is not due to increased ribosome affinity, ribosome binding site accessibility, or mRNA stability. Instead, the researchers propose a physical barrier model where the G4 structure biases ribosome movement toward the downstream start codon, thereby increasing translation output. This study provides insights into the regulatory role of 5'UTR structures in bacterial translation and highlights their potential applications in gene expression tuning.This study investigates the impact of G-quadruplex (G4) structures in the 5' untranslated region (5'UTR) of bacterial mRNA on translation efficiency. Using an in vitro translation system and E. coli, the researchers found that G4 structures in the 5'UTR enhance translation in a size-dependent manner. Longer loops within G4 structures and the presence of a hairpin upstream of the G4 further increase translation efficiency. The effect is not due to increased ribosome affinity, ribosome binding site accessibility, or mRNA stability. Instead, the researchers propose a physical barrier model where the G4 structure biases ribosome movement toward the downstream start codon, thereby increasing translation output. This study provides insights into the regulatory role of 5'UTR structures in bacterial translation and highlights their potential applications in gene expression tuning.