Preparation of a Functional GABARAP–Lipid Conjugate in Nanodiscs and its Investigation by Solution NMR Spectroscopy

Ma, Peixiang; Mohrlüder, Jeannine; Schwarten, Melanie; Stoldt, Matthias; Singh, Sameer; Hartmann, Rudolf; Pacheco, Victor; Willbold, Dieter

doi:10.1002/cbic.201000354

Cited by 24 publications

(22 citation statements)

References 65 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The best described cues are the locally enriched population of PI3P, the specific accumulation of lipidated LC3 proteins and increasingly the presence of highly curved or poorly packed membranes (Dancourt and Melia, 2014; Fan et al, 2011; Nath et al, 2014; Ragusa et al, 2012). However, neither high curvature nor PI3P are unique to autophagosomes, and while lipidated LC3 is strongly associated with autophagic processes, there is no evidence that this protein is structurally distinct from the vast pool of cytoplasmic LC3 (Ma et al, 2010); thus how proteins identify autophagosomes remains an important and intriguing question. RavZ presents one very satisfying model, that inclusion of multiple perhaps low-affinity and only moderately specific membrane recognition modules can collectively drive very specific and effective local targeting or activity of the protein.…”

Section: Resultsmentioning

confidence: 99%

The Legionella Anti-autophagy Effector RavZ Targets the Autophagosome via PI3P- and Curvature-Sensing Motifs

et al. 2015

View full text Add to dashboard Cite

Summary Autophagy is a conserved membrane transport pathway used to destroy pathogenic microbes that access the cytosol of cells. The intracellular pathogen Legionella pneumophila interferes with autophagy by delivering an effector protein, RavZ, into the host cytosol. RavZ acts by cleaving membrane-conjugated Atg8/LC3 proteins from preautophagosomal structures. Its remarkable efficiency allows minute quantities of RavZ to block autophagy throughout the cell. To understand how RavZ targets preautophagosomes and specifically acts only on membrane-associated Atg8 proteins, we elucidated its structure. Revealed is a catalytic domain related in fold to Ulp-family deubiquitinase-like enzymes and a C-terminal PI3P-binding module. RavZ targets the autophagosome via the PI3P-binding module and a catalytic domain helix and preferentially binds high-curvature membranes, intimating localization to highly curved domains in autophagosome intermediate membranes. RavZ-membrane interactions enhance substrate affinity, providing a mechanism for interfacial activation that may also be used by host autophagy proteins engaging only lipidated Atg8 proteins.

show abstract

Section: Resultsmentioning

confidence: 99%

The Legionella Anti-autophagy Effector RavZ Targets the Autophagosome via PI3P- and Curvature-Sensing Motifs

et al. 2015

View full text Add to dashboard Cite

show abstract

“…13 Detailed knowledge of the dynamics of these proteins will therefore be crucial for 6 understanding the regulation of their C-terminal modification. With molecular dynamics (MD) simulations, the time-dependent behavior and dynamics of a molecular system can be studied in silico, using conditions closely mirroring the physiological state.…”

mentioning

confidence: 99%

Conformational Polymorphism in Autophagy-Related Protein GATE-16

Schillinger

Schwarten

et al. 2015

Biochemistry

Self Cite

View full text Add to dashboard Cite

Autophagy is a fundamental homeostatic process in eukaryotic organisms, fulfilling essential roles in development and adaptation to stress. Among other factors, formation of autophagosomes critically depends on proteins of the Atg8 (autophagy-related protein 8) family, which are reversibly conjugated to membrane lipids. We have applied X-ray crystallography, nuclear magnetic resonance spectroscopy, and molecular dynamics simulations to study the conformational dynamics of Atg8-type proteins, using GATE-16 (Golgi-associated ATPase enhancer of 16 kDa), also known as GABARAPL2, as a model system. This combination of complementary approaches provides new insight into a structural transition centered on the C-terminus, which is crucial for the biological activity of these proteins.

show abstract

“…Finally, the significance of individual Bcl-2 side chains for GABARAP binding was investigated by 1 H-15 N-HSQC titration experiments using 15 N-GABARAP and Bcl-2-derived peptides. Whereas the wild-type peptide (Ser 24 -Asp 34 ) was clearly able to interact with GABARAP (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…The observation of abnormal GABARAP expression levels in cancer cells suggests a potential role of this protein during cancer development (8,9). Structural studies using nuclear magnetic resonance (NMR) spectroscopy as well as x-ray crystallography have revealed that most interactions of GABARAP are mediated by two hydrophobic pockets on its surface (10 -14), and that this binding site is also available in the lipidated form of the molecule (15). Thus, at least subsets of GABARAP binding partners are expected to compete for the interaction surface, suggesting an intricate spatio-temporal patterning of different complexes (7).…”

mentioning

confidence: 99%

Interaction of Bcl-2 with the Autophagy-related GABAA Receptor-associated Protein (GABARAP)

Schwarten

Schneider

et al. 2013

Journal of Biological Chemistry

Self Cite

View full text Add to dashboard Cite

Background: Apoptosis and autophagy are coordinately regulated, but the underlying mechanisms are incompletely understood. Results: Bcl-2 specifically interacts with GABARAP via a conserved EWD motif, resulting in impaired GABARAP lipidation. Conclusion: Sequestration of GABARAP is likely to contribute to the down-regulation of autophagy by Bcl-2. Significance: Interfering with pro-survival functions of Bcl-2 (including its impact on autophagy) represents a promising strategy for cancer therapy.

show abstract

Preparation of a Functional GABARAP–Lipid Conjugate in Nanodiscs and its Investigation by Solution NMR Spectroscopy

Cited by 24 publications

References 65 publications

The Legionella Anti-autophagy Effector RavZ Targets the Autophagosome via PI3P- and Curvature-Sensing Motifs

The Legionella Anti-autophagy Effector RavZ Targets the Autophagosome via PI3P- and Curvature-Sensing Motifs

Conformational Polymorphism in Autophagy-Related Protein GATE-16

Interaction of Bcl-2 with the Autophagy-related GABAA Receptor-associated Protein (GABARAP)

Contact Info

Product

Resources

About