Sabharwal, P. | Search bioRxiv

bioRxiv2026

Biochemical and structural characterisation of MprF homologue,LpiA from Agrobacterium

Multiple peptide resistance factor (MprF) are bi-functional enzymes encoded by several bacterial species and carry out the transfer of an amino acid from a charged tRNA to the lipid head group and further translocate the lipid across the membrane. Biochemical studies have revealed that the soluble synthase domain generates specificity and the structures of MprF have defined the general architecture of these enzymes, and that they can exist in different oligomeric states. Here, we characterise the gene product of lpiA, a MprF homologue from Agrobacterium fabrum (formerly called A. tumefaciens strain C58), a microbe that is commonly used in plant molecular biology. Cryo-EM analysis of AfMprF reveals a dimeric structure both in detergent micelle and in lipid nanodisc, and similar in architecture to the homologous enzyme from related Rhizobium sp. We further analyse some conserved residues in the soluble domain and suggest that the sulphur-aromatic motifs play a key role in substrate binding. Similar architecture of enzymes in closely related bacterial species of Agrobacterium and Rhizobium hints an evolutionary relationship but the importance of these oligomeric states in vivo remains to be analysed.

biochemistry↗

bioRxiv2026

Epistatic interaction between ribosome-associated Era GTPase and stringent response regulator RelA modulates bacterial cell growth and dormancy

Cellular homeostasis is sustained by balancing the cell growth and quiescence in response to specific environmental cues. While the mechanisms governing the transition from growth to dormancy are reasonably well understood, the processes underlying exit from dormancy remain poorly characterised. Using an integrated approach employing bacterial genetics, Cryo-EM, and molecular microbiology, we investigated this phenomenon by studying the cross-talk between two antagonistic pathways, ribosome biogenesis that promotes cell growth, and stringent response that induces dormancy. We show that the exit from dormancy requires a conserved Era GTPase whose synthesis selectively rises during onset of exponential growth phase in Escherichia coli. Era not only accelerates the maturation of head and platform regions of 30S ribosomal subunit but also dislodges RelA from 70S ribosome. This derepresses the 70S ribosome from RelA-mediated inhibition, and promotes protein synthesis driving the active cell growth. This study uncovers that temporal epistatic interaction between Era and RelA governs cellular resuscitation in bacteria. Given that the dormant bacterial populations contribute to antibiotic tolerance, understanding this regulatory axis offers new insights for resensitising the recalcitrant dormant bacteria to antibiotics.

microbiology↗