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Filaments and fingers: novel structural aspects of the single septin from Chlamydomonas reinhardtii

DOI: 10.1074/jbc.M116.762229 DOI Help

Authors: Andressa P. A. Pinto (Instituto de Fisica de Sao Carlos - USP) , Humberto M. Pereira (Instituto de Fisica de Sao Carlos - USP) , Ana E. Zeraik (Instituto de Fisica de Sao Carlos - USP) , Heloisa Ciol (Instituto de Fisica de Sao Carlos - USP) , Frederico M. Ferreira (Universidade de Santo Amaro) , Jose Brandao-neto (Diamond Light Source) , Ricardo Demarco (Instituto de Fisica de Sao Carlos - USP) , Marcos V. A. S. Navarro (Instituto de Fisica de Sao Carlos - USP) , Cristina Risi (Eastern Virginia Medical School) , Vitold E. Galkin (Eastern Virginia Medical School) , Richard C. Garratt (Instituto de Fisica de Sao Carlos - USP) , Ana P. U. Araujo (Instituto de Fisica de Sao Carlos - USP)
Co-authored by industrial partner: No

Type: Journal Paper
Journal: Journal Of Biological Chemistry

State: Published (Approved)
Published: May 2017
Diamond Proposal Number(s): 5073

Abstract: Septins are filament−forming GTP−binding proteins involved in many essential cellular events related to cytoskeletal dynamics and maintenance. Septins can self−assemble into hetero−complexes, which polymerize into highly organized, cell membrane−interacting filaments. The number of septin genes varies among organisms, and although their structure and function have been thoroughly studied in opisthokonts (including animals and fungi), no structural studies have been reported for other organisms. This makes the single septin from Chlamydomonas (CrSEPT) a particularly attractive model for investigating whether functional homopolymeric septin filaments also exist. CrSEPT was detected at the base of the flagella in Chlamydomonas, suggesting that CrSEPT is involved in the formation of a membrane diffusion barrier. Using transmission electron microscopy, we observed that recombinant CrSEPT forms long filaments with dimensions comparable to those of the canonical structure described for opisthokonts. The GTP−binding domain of CrSEPT purified as a nucleotide−free monomer that hydrolyzes GTP and readily binds its analog GTPγS. We also found that on nucleotide binding, CrSEPT forms dimers that are stabilized by an interface involving the ligand (G−interface). Across this interface, one monomer supplied a catalytic arginine to the opposing subunit, greatly accelerating the rate of GTP hydrolysis. This is the first report of an arginine finger observed in a septin and suggests that CrSEPT may act as its own GTP−activating protein (GAP). The finger is conserved in all algal septin sequences, suggesting a possible correlation between the ability to form homo−polymeric filaments and the accelerated rate of hydrolysis which it provides.

Journal Keywords: algae; arginine finger; Chlamydomonas; crystal structure; GTPase; filament; septin

Subject Areas: Biology and Bio-materials

Instruments: I04-1-Macromolecular Crystallography (fixed wavelength)