Crystal Structure of a Schistosoma mansoni Septin Reveals the Phenomenon of Strand Slippage in Septins Dependent on the Nature of the Bound Nucleotide

Zeraik, Ana Eliza; Pereira, H.M.; Santos, Y. Volpato; Brandão-Neto, J.; Spoerner, Michael; Santos, Maiara S.; Colnago, Luiz Alberto; Garratt, R.C.; Araújo, Ana Paula Ulian de; DeMarco, Ricardo

doi:10.1074/jbc.m113.525352

Cited by 40 publications

(53 citation statements)

References 55 publications

(51 reference statements)

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“…The structures of SmSEPT10 and human SEPT2 also present elongated strands, but only when bound to either GTP or one of its analogues, as is the case for CrSEPT. In SmSEPT10, the ␤3 strand is known to undergo slippage as a function of the nature of nucleotide (GDP or GTP) present, and the hydrogenbonding pattern observed here for CrSEPT is entirely consistent with the GTP-bound state (24).…”

Section: Overall Structure Of Crsept Gtp-binding Domainsupporting

confidence: 49%

“…Other variations at the NC interface have been observed in the crystal structures of SmSEPT10 and human SEPT3 (20,24). In the former, ␤-strand slippage as a function of the nature of the bound nucleotide leads to strand projection into the interface, whereas in the latter, a dramatic rearrangement of the salt bridges causes a squeezing of the interface in which the centers of mass of the two subunits lie closer together.…”

Section: Structure Of C Reinhardtii Septinmentioning

confidence: 93%

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

Pinto

Pereira

Zeraik

et al. 2017

Journal of Biological Chemistry

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Edited by Norma AllewellSeptins are filament-forming GTP-binding proteins involved in many essential cellular events related to cytoskeletal dynamics and maintenance. Septins can self-assemble into heterocomplexes, which polymerize into highly organized, cell membraneinteracting 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 with 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 guanosine 5-3-O-(thio)triphosphate. We also found that upon nucleotide binding, CrSEPT formed dimers that were 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. The finger is conserved in all algal septin sequences, suggesting a possible correlation between the ability to form homopolymeric filaments and the accelerated rate of hydrolysis that it provides.

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Section: Overall Structure Of Crsept Gtp-binding Domainsupporting

confidence: 49%

Section: Structure Of C Reinhardtii Septinmentioning

confidence: 93%

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

Pinto

Pereira

Zeraik

et al. 2017

Journal of Biological Chemistry

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“…SmSEPT10 is unable to hydrolyze GTP, as we have previously shown [27], although it can bind both GTP and GDP. This is consistent with SmSEPT10 belonging to the SEPT6 subgroup, which is known to lack GTPase activity.…”

Section: Discussionmentioning

confidence: 87%

“…A preliminary characterization of nucleotide binding for SmSEPT10, a catalytically inactive protein, has been previously performed [27] and therefore this aspect was only explored for SmSEPT5. The nucleotide content of the purified SmSEPT5 was analyzed by anion exchange chromatography and no bound 7 nucleotide was detected after the second round of purification, suggesting a low affinity for the nucleotide (not shown) and justifying the appropriateness of the purified proteins in ITC and GTP hydrolysis experiments.…”

Section: Nucleotide Binding and Hydrolysis By Smsept5mentioning

confidence: 99%

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Biophysical dissection of schistosome septins: Insights into oligomerization and membrane binding

et al. 2016

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Citation for published item:erikD eF nd tykovD wF nd pontesD wF nd xemuritD sF nd uinlnD F nd er¡ ujoD eF F nd hewroD F @PHITA 9fiophysil dissetion of shistosome septins X insights into oligomeriztion nd memrne indingF9D fiohimieFD IQI F ppF WTEIHSF Further information on publisher's website: Use policyThe full-text may be used and/or reproduced, and given to third parties in any format or medium, without prior permission or charge, for personal research or study, educational, or not-for-prot purposes provided that:• a full bibliographic reference is made to the original source • a link is made to the metadata record in DRO • the full-text is not changed in any way The full-text must not be sold in any format or medium without the formal permission of the copyright holders.Please consult the full DRO policy for further details. the N-and C-terminal interfaces in a nucleotide independent fashion but form heterodimers via the G interface, which are nucleotide dependent. Furthermore, we report for the first time that it is the C-terminus of SmSETP10, rather than the Nterminal polybasic region found in other septins, that mediates its binding to liposomes.Upon binding we observe formation of discrete lipo-protein clusters and higher order septin structures, making our system an exciting model to study interactions of septins with biological membranes.2

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Lean forward: Genetic analysis of temperature‐sensitive mutants unfolds the secrets of oligomeric protein complex assembly

McMurray

2014

BioEssays

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Multisubunit protein complexes are essential for cellular function. Genetic analysis of essential processes requires special tools, among which temperature-sensitive (Ts) mutants have historically been crucial. Many researchers assume that the effect of temperature on such mutants is to drive their proteolytic destruction. In fact, degradation-mediated elimination of mutant proteins likely explains only a fraction of the phenotypes associated with Ts mutants. Here I discuss insights gained from analysis of Ts mutants in oligomeric proteins, with particular focus on the study of septins, GTP-binding subunits of cytoskeletal filaments whose structures and functions are the subject of current investigation in my and many other labs. I argue that the kinds of unbiased forward genetic approaches that generate Ts mutants provide information that is largely inaccessible to modern reverse genetic methodologies, and will continue to drive our understanding of higher-order assembly by septins and other oligomeric proteins.

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Crystal Structure of a Schistosoma mansoni Septin Reveals the Phenomenon of Strand Slippage in Septins Dependent on the Nature of the Bound Nucleotide

Cited by 40 publications

References 55 publications

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

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

Biophysical dissection of schistosome septins: Insights into oligomerization and membrane binding

Lean forward: Genetic analysis of temperature‐sensitive mutants unfolds the secrets of oligomeric protein complex assembly

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