Mg2+ binding triggers rearrangement of the IM30 ring structure, resulting in augmented exposure of hydrophobic surfaces competent for membrane binding

Heidrich, Jennifer; Junglas, Benedikt; Grytsyk, Natalia; Hellmann, Nadja; Rusitzka, Kristiane A. K.; Gebauer, Wolfgang; Markl, Jürgen; Hellwig, Petra; Schneider, Dirk

doi:10.1074/jbc.ra117.000991

Cited by 17 publications

(36 citation statements)

References 59 publications

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“…Furthermore, the contribution of elastic and viscous behavior might shift in this process. The viscoelastic properties observed in presence of Mg 2+ , which stabilizes the ring structures (35), are more similar to the viscoelastic properties observed in the initial phase of IM30 membrane-interaction in the absence of Mg 2+ (Fig. 3C).…”

Section: Mg 2+ Binding Alters the Viscoelastic Properties Of Im30 Decsupporting

confidence: 76%

“…Yet, IM30 has also been shown to mediate membrane fusion, at least in vitro, which involves membrane destabilization. However, as shown recently, the presence of Mg 2+ is required for IM30 acting as a membrane fusion protein (10,35). Binding of IM30 to negatively charged membranes has been observed using small PG liposomes, PC/PG GUVs, and PG SLBs in the absence of Mg 2+ (Fig.…”

Section: Discussionsupporting

confidence: 59%

“…Hence, the particle diameters differ significantly in absence vs presence of Mg 2+ . In fact, in presence of Mg 2+ IM30 rings are stabilized and increased ring-stacking and formation of double rings was observed in solution (35). The increased height of membrane-attached IM30 in presence of Mg 2+ likely originates from stabilization of IM30 supercomplexes and membrane attachment of higher-ordered IM30 structures and explains the higher viscoelasticity observed in the QCM measurements ( Fig.…”

Section: Mg 2+ Triggers Im30 Induced Formation Of Bilayer Defectsmentioning

confidence: 82%

“…Binding of Mg 2+ to IM30 alters the structure of IM30 and triggers membrane fusion (10,35). Due to the exposure of an increased hydrophobic surface upon Mg 2+ -binding (35), the structural changes induced by Mg 2+ -binding potentially enhance its membrane binding affinity. To test this assumption, we first assessed whether binding of IM30 to native TMs depends on the cytosolic Mg 2+ concentrations.…”

Section: Mg 2+ Increases the Membrane-binding Propensity Of Im30mentioning

confidence: 99%

“…Once Mg 2+ has entered the chloroplast stroma or cyanobacterial cytoplasm, respectively, out of the thylakoid lumen, it allosterically regulates the activity of several enzymes and proteins (33,34). Indeed, Mg 2+ directly binds also to IM30 in solution, inducing several structural rearrangements, resulting in an increased surface hydrophobicity and increased protease resistance of the protein´s C-terminal domain (35). Thus, IM30triggered membrane remodeling is potentially controlled via Mg 2+ -binding to IM30.…”

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confidence: 99%

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Mg²⁺-binding shifts the IM30 activity from membrane protection to membrane destabilization

Junglas

Axt

Siebenaller

et al. 2020

Preprint

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The inner membrane-associated protein of 30 kDa (IM30) is essential in chloroplasts and cyanobacteria. The spatio-temporal cellular localization of the protein appears to be highly dynamic and triggered by internal as well as external stimuli, mainly light intensity. A soluble fraction of the protein is localized in the cyanobacterial cytoplasm or the chloroplast stroma, respectively. Additionally, the protein attaches to the thylakoid membrane as well as to the chloroplast inner envelope or the cyanobacterial cytoplasmic membrane, respectively, especially under conditions of membrane stress. IM30 is involved in thylakoid membrane biogenesis and/or maintenance, where it either stabilizes membranes and/or triggers membrane-fusion processes. These apparently contradicting processes have to be tightly controlled and separated spatiotemporally in chloroplasts and cyanobacteria. The latter process depends on Mg2+-binding to IM30; yet, it still is unclear how Mg2+-loaded IM30 interacts with membranes and promotes membrane fusion. Here we show that interaction of Mg2+ with IM30 results in increased binding of IM30 to native as well as model membranes. Via Atomic Force Microscopy in liquid, IM30-induced bilayer defects were observed in solid-supported bilayers in presence of Mg2+. The observed interaction of IM30 with membrane surfaces differs dramatically from previously observed membrane-stabilizing, carpet-like structures in the absence of Mg2+. Mg2+-induced alterations of the IM30 structure switches the IM30 activity from a membrane-stabilizing to a membrane-destabilizing function, a crucial step in membrane fusion.

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Section: Mg 2+ Binding Alters the Viscoelastic Properties Of Im30 Decsupporting

confidence: 76%

Section: Discussionsupporting

confidence: 59%

Section: Mg 2+ Triggers Im30 Induced Formation Of Bilayer Defectsmentioning

confidence: 82%

Section: Mg 2+ Increases the Membrane-binding Propensity Of Im30mentioning

confidence: 99%

mentioning

confidence: 99%

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Mg²⁺-binding shifts the IM30 activity from membrane protection to membrane destabilization

Junglas

Axt

Siebenaller

et al. 2020

Preprint

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Binding and/or hydrolysis of purine‐based nucleotides is not required for IM30 ring formation

et al. 2021

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IM30, the inner membrane-associated protein of 30 kDa, is conserved in cyanobacteria and chloroplasts. Although its exact physiological function is still mysterious, IM30 is clearly essential for thylakoid membrane biogenesis and/or dynamics. Recently, a cryptic IM30 GTPase activity has been reported, albeit thus far no physiological function has been attributed to this. Yet, it is still possible that GTP binding/hydrolysis affects formation of the prototypical large homo-oligomeric IM30 ring and rod structures. Here, we show that the Synechocystis sp. PCC 6803 IM30 protein in fact is an NTPase that hydrolyzes GTP and ATP, but not CTP or UTP, with about identical rates. While IM30 forms large oligomeric ring complexes, nucleotide binding and/or hydrolysis are clearly not required for ring formation.

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Monomer unfolding of a bacterial ESCRT‐III superfamily member is coupled to oligomer disassembly

Quarta,

Bhandari,

Girard

et al. 2024

Protein Science

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The inner membrane associated protein of 30 kDa (IM30), a member of the endosomal sorting complex required for transport (ESCRT‐III) superfamily, is crucially involved in the biogenesis and maintenance of thylakoid membranes in cyanobacteria and chloroplasts. In solution, IM30 assembles into various large oligomeric barrel‐ or tube‐like structures, whereas upon membrane binding it forms large, flat carpet structures. Dynamic localization of the protein in solution, to membranes and changes of the oligomeric states are crucial for its in vivo function. ESCRT‐III proteins are known to form oligomeric structures that are dynamically assembled from monomeric/smaller oligomeric proteins, and thus these smaller building blocks must be assembled sequentially in a highly orchestrated manner, a still poorly understood process. The impact of IM30 oligomerization on function remains difficult to study due to its high intrinsic tendency to homo‐oligomerize. Here, we used molecular dynamics simulations to investigate the stability of individual helices in IM30 and identified unstable regions that may provide structural flexibility. Urea‐mediated disassembly of the IM30 barrel structures was spectroscopically monitored, as well as changes in the protein's tertiary and secondary structure. The experimental data were finally compared to a three‐state model that describes oligomer disassembly and monomer unfolding. In this study, we identified a highly stable conserved structural core of ESCRT‐III proteins and discuss the advantages of having flexible intermediate structures and their putative relevance for ESCRT‐III proteins.

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Mg2+ binding triggers rearrangement of the IM30 ring structure, resulting in augmented exposure of hydrophobic surfaces competent for membrane binding

Cited by 17 publications

References 59 publications

Mg²⁺-binding shifts the IM30 activity from membrane protection to membrane destabilization

Mg²⁺-binding shifts the IM30 activity from membrane protection to membrane destabilization

Binding and/or hydrolysis of purine‐based nucleotides is not required for IM30 ring formation

Monomer unfolding of a bacterial ESCRT‐III superfamily member is coupled to oligomer disassembly

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Mg2+ binding triggers rearrangement of the IM30 ring structure, resulting in augmented exposure of hydrophobic surfaces competent for membrane binding

Cited by 17 publications

References 59 publications

Mg2+-binding shifts the IM30 activity from membrane protection to membrane destabilization

Mg2+-binding shifts the IM30 activity from membrane protection to membrane destabilization

Binding and/or hydrolysis of purine‐based nucleotides is not required for IM30 ring formation

Monomer unfolding of a bacterial ESCRT‐III superfamily member is coupled to oligomer disassembly

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Mg²⁺-binding shifts the IM30 activity from membrane protection to membrane destabilization

Mg²⁺-binding shifts the IM30 activity from membrane protection to membrane destabilization