Biology subjects

Grupp, B.

Publications and source records attributed to Grupp, B..

4 recordsLinked to original sources

bioRxiv2026

A nucleotide-triggered molecular switch orchestrating septin polymerization

Septins are conserved cytoskeletal GTP-binding proteins that form higher-order structures critical for cytokinesis and polarized growth across opisthokonts. In budding yeast and humans, four homologous septins assemble into hetero-octameric protofilaments through alternating interactions between adjacent G-domains (G-interface) or N- and C-termini (NC-interface), which then polymerize end-to-end into filaments. How nucleotide binding and hydrolysis control filament assembly has remained elusive. We uncover a septin-specific mechanism in which nucleotide binding stabilizes the monomeric G-interface and primes it for protofilament formation. Cryo-EM and DEER spectroscopy reveal nucleotide-induced conformational changes that engage the G-interface and enable an NC-compatible conformation absent in the nucleotide-free state. In vitro binding and reconstitution assays confirm that G-interface formation is strictly nucleotide-dependent and required for subsequent NC-interface assembly. These findings establish unidirectional allosteric signaling from the G-to the NC-interface, revealing the molecular basis for controlled septin protofilament assembly.

molecular biology↗

bioRxiv2025

Interface integrity in septin protofilaments is maintained by an arginine residue conserved from yeast to man.

The septins are conserved, filament-forming, guanine nucleotide binding cytoskeletal proteins. They assemble into palindromic protofilaments which polymerize further into higher-ordered structures that participate in essential intracellular processes such as cytokinesis or polarity establishment. Septins belong structurally to the P-Loop NTPases but, unlike their relatives Ras or Rho, do not mediate signals to effectors through GTP binding and hydrolysis. Biochemical approaches addressing how and why septins utilize nucleotides are hampered by the lack of nucleotide free complexes. Using molecular dynamics simulations, we determined structural alterations and inter-subunit binding free energies in human and yeast septin dimer structures and in their in silico generated apo forms. An interchain saltbridge network in the septin unique {beta}-meander, conserved across all kingdoms of septin containing species, is destabilized upon nucleotide removal, concomitant with disruption of the entire G-interface. Within this network, we identified a conserved arginine residue, Arg({beta}b), as the central interaction hub. The essential role of Arg({beta}b) for interface integrity was confirmed species-overlapping in human and yeast septins by in vitro and in vivo experimentation.

biochemistry↗

bioRxiv2023

Transient septin sumoylation steers a Fir1-Skt5 protein complex between the split septin ring

Ubiquitylation and phosphorylation control composition and architecture of the cell separation machinery in yeast and other eukaryotes. The significance of septin sumoylation on cell separation remained an enigma. Septins form an hourglass structure at the bud neck of yeast cells that transforms into a split septin double ring during mitosis. We discovered that sumoylated septins recruit the cytokinesis checkpoint protein Fir1 to the peripheral side of the septin hourglass. Subsequent de-sumoylation and synchronized binding to the scaffold Spa2 relocate Fir1 in a seamless transition between the split septin rings. Fir1 binds and carries Skt5 on its route to the division plane where the Fir1-Skt5 complex serves as receptor for chitin synthase III. We propose that the opposite positioning of the sumoylated septins and Spa2 creates a tension across the ring that upon de-sumoylation tunnels the membrane-bound Fir1-Skt5 complex through a transiently permeable septin diffusion barrier.

cell biology↗

bioRxiv2022

An in silico approach to determine inter-subunit affinities in human septin complexes

The septins are a conserved family of filament-forming guanine nucleotide binding proteins, often named the fourth component of the cytoskeleton. Correctly assembled septin structures are required for essential intracellular processes such as cytokinesis, vesicular transport, polarity establishment, and cellular adhesion. Structurally, septins belong to the P-Loop NTPases but they do not mediate signals to effectors through GTP binding and hydrolysis. GTP binding and hydrolysis are believed to contribute to septin complex integrity, but biochemical approaches addressing this topic are hampered by the stability of septin complexes after recombinant expression and the lack of nucleotide-depleted complexes. To overcome this limitation, we used a molecular dynamics-based approach to determine inter-subunit binding free energies in available human septin dimer structures and in their apo forms, which we generated in silico. The nucleotide in the GTPase active subunits SEPT2 and SEPT7, but not in SEPT6, was identified as a stabilizing element in the G interface as it is coordinated at its ribose ring to conserved amino acids. Removal of GDP from SEPT2 and SEPT7 results in flipping of a conserved Arg residue and disruption of an extensive hydrogen bond network in the septin unique element, concomitant with a decreased inter-subunit affinity.

biochemistry↗