On the role of nucleotides and lipids in the polymerization of the actin homolog MreB from a Gram-positive bacterium

Abstract

In vivo, bacterial actin MreB assembles into dynamic membrane-associated filamentous structures that exhibit circumferential motion around the cell. Current knowledge of MreB biochemical and polymerization properties in vitro remains limited and is mostly based on MreB proteins from Gram-negative species. In this study, we report the first observation of organized protofilaments by electron microscopy and the first 3D-structure of MreB from a Gram-positive bacterium. We show that Geobacillus stearothermophilus MreB forms straight pairs of protofilaments on lipid surfaces in the presence of ATP or GTP, but not in the presence of ADP, GDP or non-hydrolysable ATP analogs. We demonstrate that membrane anchoring is mediated by two spatially close short hydrophobic sequences while electrostatic interactions also contribute to lipid binding, and show that the population of membrane-bound protofilament doublets is in steady-state. In solution, protofilament doublets were not detected in any condition tested. Instead, MreB formed large sheets regardless of the bound nucleotide, albeit at a higher critical concentration. Altogether, our results indicate that both lipids and ATP are facilitators of MreB polymerization, and are consistent with a dual effect of ATP hydrolysis, in promoting both membrane binding and filaments assembly/disassembly.

Alanine-scanning mutagenesis library of MreB reveals distinct roles for regulating cell shape and viability

“…The actin superfamily is defined by conserved structural motifs needed for ATP binding/hydrolysis and polymerization. MreB has all of the structural features one would expect in an actin, including the ability to bind ATP with the nucleotide binding pocket or cleft between domains I and II 28,41,71,80,81 .…”

Probing the Molecular Interactions of A22 with Prokaryotic Actin MreB and Eukaryotic Actin: A Computational and Experimental Study

“…MreB polymerizes in the presence of ATP or GTP, forming double filaments assembled in an antiparallel orientation. In the ATP-bound state, MreB undergoes conformational rearrangement, facilitating nucleotide hydrolysis and depolymerisation [11][12][13] . Although the role of hydrolysis in polymerization dynamics remains unclear, the dynamic organization of the MreB filaments is well understood to mediate cell wall synthesis and cell shape maintenance in bacterial cells 9,11,14 .…”

Getting bacterial cells into shape

“…Now, in eLife, Rut Carballido-López and colleagues at research institutes in France and Germany – including Wei Mao (Université Paris-Saclay) as first author – report new insights into the dynamics of MreB in the bacterium Geobacillus stearothermophilus ( Mao et al, 2023 ). Mao et al used a combination of techniques to study the structure of MreB filaments and how they assemble and disassemble.…”