NMR- and Circular Dichroism-monitored Lipid Binding Studies Suggest a General Role for the FATC Domain as Membrane Anchor of Phosphatidylinositol 3-Kinase-related Kinases (PIKK)

Sommer, Lisa A. M.; Schaad, Martin; Dames, Sonja A.

doi:10.1074/jbc.m113.467233

Cited by 18 publications

(36 citation statements)

References 83 publications

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“…Some of the mutants not binding to liposomes may be incorporated in full-length TOR for in vivo functional and localization studies, which will help to better understand the role of the redox-sensitive FATC domain for the regulation of TOR localization at different cellular membranes and its local signaling activity. Recent studies suggest that the ability to interact with membrane mimetics is also shared by the FATC domains of the other PIKKs [24]. The presented binding data of different TOR FATC mutants it also helpful to better rationalize, why the FATC domains of all tested PIKKs can interact with membrane mimetics, although with apparently somehow different preferences for different membrane properties such as surface charge, shape, and curvature and the acyl chain packing density.…”

Section: Discussionmentioning

confidence: 99%

“…2-4, SI Figs. S3-S5) as well as from previous studies indicated that the B1 domain of streptococcal protein G (GB1), alone or fused to other proteins, does not interact with DPC micelles, DihepPC/DMPC bicelles, or SUVs [24,39]. Recently, we further characterized the interaction of the N-terminal domain of Formin C, of a 26-mer peptide corresponding to a large unstructured loop of this domain, and of a mutant form lacking this loop with different membrane mimetics.…”

Section: Discussionmentioning

confidence: 99%

“…Another example for a protein region that shows no interactions with DPC micelles is the N-terminal natively unstructured region of the mycobacterial kinase G (PknG, residues 1-76, Uniprot ID P65728) (unpublished data). Finally, two more examples for proteins that show as all tested TOR FATC mutants strong spectral changes with micelles and bicelles but only weak ones with SUVs are the FRB domain of TOR [34] and the FATC domain of the PIKK DNA-PKcs [24]. Generally, the different affinities for different membrane mimetics can often be rationalized based on the current knowledge of the protein function.…”

Section: Discussionmentioning

confidence: 99%

“…Based on recent NMR-and CD-monitored binding studies, the FATC domains of the other human PIKK family members including ataxia-telangiectasia mutated (ATM), ataxia-and Rad3-related (ATR), DNA-dependent protein kinase catalytic subunit (DNA-PKcs), suppressor of morphogenesis in genitalia (SMG-1), and transformation/transcription domain-associated protein (TRRAP) can also interact with membrane mimetics, which results generally in an increase of the a-helical secondary structure content [24].…”

Section: Introductionmentioning

confidence: 99%

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Characterization of residue‐dependent differences in the peripheral membrane association of the FATC domain of the kinase ‘target of rapamycin’ by NMR and CD spectroscopy

Sommer¹,

Dames²

2014

FEBS Letters

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Edited by Sandro SonninoThe conserved C-terminal FATC domain of the kinase 'target of rapamycin' is important for its regulation and was suggested to contain a peripheral membrane anchor. Here, we present the characterization of the interactions of the yeast TOR1 FATC domain (2438-2470 = y1fatc) and 15 mutants with membrane mimetic micelles, bicelles, and small unilamellar vesicles (SUVs) by NMR and CD spectroscopy. Replacement of up to 6-7 residues did not result in a significant abrogation of the association with micelles or bicelles. However, replacement of only one residue could result in an impairment of the interaction with SUVs that are usually used at low concentrations. Some mutants not binding liposomes may be introduced in full-length TOR for future functional and localization studies in vivo.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Characterization of residue‐dependent differences in the peripheral membrane association of the FATC domain of the kinase ‘target of rapamycin’ by NMR and CD spectroscopy

Sommer¹,

Dames²

2014

FEBS Letters

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“…Many peripheral proteins, such as myristoylated alanine-rich C kinase substrate (MARCKS), 10 FAT C-terminal (FATC) domain, 11 and LL37, 12 contain clusters of basic and aromatic residues that interact with lipid bilayers. Generally, aromatic and basic residue motifs are important for the anchoring of proteins to the biomembranes, 13 particularly aromatic residues, which may play key roles as efficient anchors during binding at the phospholipid membrane interface.…”

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confidence: 99%

The chain order of binary unsaturated lipid bilayers modulated by aromatic-residue-containing peptides: an ATR-FTIR spectroscopy study

et al. 2017

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In this study, we described the chain order change of binary unsaturated lipid bilayers by studying pure 2-oleoyl-1-pamlitoyl-sn-glyecro-3-phosphocholine (POPC) bilayers, 2-oleoyl-1-pamlitoyl-sn-glyecro-3-glycerol (POPG) bilayers, and POPC/POPG bilayers. The interactions between the POPC/POPG membrane and three model peptides containing basic, aromatic, and hydrophobic residues were studied to investigate the effects of the peptide orientation on the acyl chain order of the POPC/POPG bilayers.Attenuated total reflectance-Fourier transform infrared (ATR-FTIR) spectroscopic studies showed that the acyl chain order of the POPC/POPG bilayers decreased as compared to that of the pure POPC or POPG bilayers. The chain order of the POPC/POPG bilayers significantly decreased when Gravin was added as compared to that when Vamp2 or Antp was added. Fluorescence spectroscopic measurements further show that the tryptophan of the three peptides may interact with the hydrophobic region of the POPC/POPG bilayers, thus resulting in a change in the conformational order of the POPC/POPG acyl chains. This study may provide valuable insight into the dynamics and flexibility of lipid bilayers with different lipids and highlights the importance of aromatic residues in peptide binding to the membrane.

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Target of rapamycin FATC domain as a general membrane anchor: The FKBP‐12 like domain of FKBP38 as a case study

2017

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Increased efforts have been undertaken to better understand the formation of signaling complexes at cellular membranes. Since the preparation of proteins containing a transmembrane domain or a prenylation motif is generally challenging an alternative membrane anchoring unit that is easy to attach, water-soluble and binds to different membrane mimetics would find broad application. The 33-residue long FATC domain of yeast TOR1 (y1fatc) fulfills these criteria and binds to neutral and negatively charged micelles, bicelles, and liposomes. As a case study, we fused it to the FKBP506-binding region of the protein FKBP38 (FKBP38-BD) and used H- N NMR spectroscopy to characterize localization of the chimeric protein to micelles, bicelles, and liposomes. Based on these and published data for y1fatc, its use as a C-terminally attachable membrane anchor for other proteins is compatible with a wide range of buffer conditions (pH circa 6-8.5, NaCl 0 to >150 mM, presence of reducing agents, different salts such as MgCl and CaCl ). The high water-solubility of y1fatc enables its use for titration experiments against a membrane-localized interaction partner of the fused target protein. Results from studies with peptides corresponding to the C-terminal 17-11 residues of the 33-residue long domain by 1D H NMR and CD spectroscopy indicate that they still can interact with membrane mimetics. Thus, they may be used as membrane anchors if the full y1fatc sequence is disturbing or if a chemically synthesized y1fatc peptide shall be attached by native chemical ligation, for example, unlabeled peptide to N-labeled target protein for NMR studies.

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NMR- and Circular Dichroism-monitored Lipid Binding Studies Suggest a General Role for the FATC Domain as Membrane Anchor of Phosphatidylinositol 3-Kinase-related Kinases (PIKK)

Cited by 18 publications

References 83 publications

Characterization of residue‐dependent differences in the peripheral membrane association of the FATC domain of the kinase ‘target of rapamycin’ by NMR and CD spectroscopy

Characterization of residue‐dependent differences in the peripheral membrane association of the FATC domain of the kinase ‘target of rapamycin’ by NMR and CD spectroscopy

The chain order of binary unsaturated lipid bilayers modulated by aromatic-residue-containing peptides: an ATR-FTIR spectroscopy study

Target of rapamycin FATC domain as a general membrane anchor: The FKBP‐12 like domain of FKBP38 as a case study

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