FEBS Letters 583 (2009) 2854–2858

journal homepage: www.FEBSLetters.org

Ganglioside-induced amyloid formation by human islet amyloid polypeptide

in lipid rafts
Masaki Wakabayashi 1, Katsumi Matsuzaki *
Graduate School of Pharmaceutical Sciences, Kyoto University, Sakyo-ku, Kyoto 606-8501, Japan

a r t i c l e i n f o a b s t r a c t

Article history: Human islet amyloid polypeptide (hIAPP) is the primary component of the amyloid deposits found
Received 17 June 2009 in the pancreatic islets of patients with type 2 diabetes mellitus. However, it is unknown how amy-
Revised 20 July 2009 loid fibrils are formed in vivo. In this study, we demonstrate that gangliosides play an essential role
Accepted 24 July 2009
in the formation of amyloid deposits by hIAPP on plasma membranes. Amyloid fibrils accumulated
Available online 3 August 2009
in ganglioside- and cholesterol-rich microscopic domains (‘lipid rafts’). The depletion of ganglio-
Edited by Jesus Avila sides or cholesterol significantly reduced the amount of amyloid deposited. These results clearly
showed that the formation of amyloid fibrils was mediated by gangliosides in lipid rafts.
Keywords:
Ó 2009 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
Human islet amyloid polypeptide
Amyloid fibril
Ganglioside
Cholesterol
Lipid raft

1. Introduction
The deposition of insoluble amyloid fibrils in tissues is a hall-
mark of a range of diseases referred to as amyloidoses, that include
type II diabetes mellitus (DM), Alzheimer’s disease, and transmis-
sible spongiform encephalopathy [1–4]. The misfolding of a unique
peptide or protein into b-sheet-rich fibrous structures is proposed
to be a key step in the onset and development of these diseases
[3,5]. The amyloid deposits cause cell dysfunction, death, and sub-
sequently severe impairment in tissues of patients. Elucidation of
amyloid formation mechanisms, which are still poorly understood,
is essential for prevention of the onset and development of
amyloidoses.
Native human islet amyloid polypeptide (hIAPP) is the primary
component of the amyloid deposits [6,7] found in the pancreatic is-
let of patients with type 2 DM [8,9]. The causal relationship be-
tween the formation of amyloid fibrils containing hIAPP and the
Abbreviations: aMEM, alpha modification of Eagle’s medium; CHO, Chinese
hamster ovary; CTX-B, Alexa Fluor 647-conjugated cholera toxin subunit B; DIC,
differential interference contrast; DM, diabetes mellitus; DMEM, Dulbecco’s Mod-
ified Eagle’s Medium; HFIP, hexafluoroisopropanol; hIAPP, human islet amyloid
polypeptide; PDMP, N-[2-hydroxy-1-(4-morpholinylmethyl)-2-phenylethyl]-
decanamide hydrochloride; TBS, Tris buffered saline
* Corresponding author. Fax: +81 75 753 4578.
E-mail address: katsumim@pharm.kyoto-u.ac.jp (K. Matsuzaki).
1
Research fellow of the Japan Society for the Promotion of Science.
onset of type 2 DM has not been resolved. However, the amyloid
deposits cause cellular dysfunction, suggesting that their forma-
tion is involved in the pathological process. It should be noted that
the physiological concentration of hIAPP might not be high enough
to cause self-aggregation. Accumulating evidence has suggested
that membrane lipids play a crucial role in the formation of amy-
loid fibrils [10]. Experiments in vitro demonstrated that two nega-
tively charged lipid species, phosphatidylglycerol (PG) [11], and
phosphatidylserine (PS) [12], significantly facilitate the production
of amyloid by hIAPP. However, neither lipid is expressed in the
outer leaflets of plasma membranes of mammalian cells. Therefore,
other lipid species would induce the amyloidogenesis on cell mem-
branes. In the case of Alzheimer’s amyloid b-protein, gangliosides
in lipid microdomains (‘lipid rafts’) mediated the protein’s aggrega-
tion [13]. Lipid rafts are composed of sphingomyelin, gangliosides,
and cholesterol [14] and considered to be a platform of a large
number of events in plasma membranes, including signal trans-
ductions and viral assembly and budding [15–17]. The formation
of amyloid fibrils by hIAPP is also possibly mediated by ganglio-
sides in lipid rafts on live cell membranes.
In this study, we visualized hIAPP amyloid deposits on cell
membranes. Amyloid fibrils accumulated in ganglioside- and cho-
lesterol-rich domains in a concentration-dependent manner, lead-
ing to cell death. The depletion of gangliosides or cholesterol
significantly reduced the amount of hIAPP accumulated.

0014-5793/$36.00 Ó 2009 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
doi:10.1016/j.febslet.2009.07.044
 M. Wakabayashi, K. Matsuzaki / FEBS Letters 583 (2009) 2854–2858
2. Materials and methods
2.1. Materials
Dulbecco’s Modified Eagle’s Medium (DMEM), horse serum, bo-
vine serum, penicillin, and streptomycin were purchased from
Invitrogen (Carlsbad, CA). Alpha modification of Eagle’s medium
(aMEM) and Congo red were obtained from ICN Biomedicals (Aur-
ora, OH) and Nacalai Tesque (Kyoto, Japan), respectively. HIAPP
(purity > 95%) was purchased from Peptide Institute (Minou, Ja-
pan). Alexa Fluor 647-conjugated cholera toxin subunit B (CTX-B)
and Calcein-AM were obtained from Molecular Probes (Eugene,
OR). Hexafluoroisopropanol (HFIP) was purchased from Sigma
(St. Louis, MO). N-[2-Hydroxy-1-(4-morpholinylmethyl)-2-phenyl-
ethyl]-decanamide hydrochloride (PDMP) and compactin (meva-
statin) were obtained from Wako (Osaka, Japan) and Pierce
(Rockford, IL), respectively.
2.2. Peptide solutions
In order to prepare stock solutions of aggregate-free hIAPP, the
peptide was first dissolved in HFIP at 500 lM and aliquots of the
solution were nitrogen-evaporated to completely remove the sol-
vent. Peptide stock solutions were stored at 80 °C prior to use.
Just before the experiment, hIAPP was dissolved in 10 mM sodium
phosphate buffer and pH was adjusted to pH 10 with 1 M NaOH
[18].
To confirm the absence of any pre-aggregated peptide, prepared
stock solution of hIAPP was fractionated with a Superdex 75 10/
300 GL column with an exclusion limit of 40 000 Da, which had
been pretreated with an excess of BSA to block non-specific bind-
ing of peptides. The column was eluted with Tris buffered saline
(TBS) at a flow rate of 0.5 mL/min and the peptide was detected
by measuring UV absorbance at 220 nm. Molecular mass was esti-
mated with FITC-dextrans (M.W. 4400, 21 200, 42 000) as stan-
dards. The blank chromatographs were subtracted. From the
prepared solution, any oligomers and aggregates were not detected
(data not shown).
2.3. Cell culture and sample preparation
Rat pheochromocytoma PC12 cells were cultured in DMEM con-
taining 5% horse serum, 10% bovine serum, 100 units/mL penicillin,
and 0.1 mg/mL of streptomycin at 37 °C with 5% CO2. Chinese ham-
ster ovary (CHO) cells were cultured in aMEM containing 10% fetal
bovine serum, 100 U/mL penicillin, and 0.1 mg/mL of streptomycin
at 37 °C with 5% CO2. After being plated at a density of 100 000
cells onto a poly-D-lysine-coated 35-mm glass-bottomed dish, cells
were incubated for 24 h at 37 °C with 5% CO2. Unless otherwise
noted, PC12 and CHO cells were incubated with reagents in the
medium at 37 °C with 5% CO2 and washed twice with TBS. The cells
were stained with fluorophores in TBS at room temperature and
washed twice with TBS.
2.4. Cell staining
Cells were incubated with freshly prepared hIAPP and subse-
quently stained with 20 lM Congo red (excitation, 543 nm; emis-
sion, band pass filter (BP) 560–615 nm) for 30 min. Labeling of
cell surface gangliosides was performed by incubating the cells
with 10 lg/mL CTX-B (excitation, 633 nm; emission, LP 650 nm)
for 20 min. Fluorescently labeled cells were visualized using the
63 C-Apochromat objective of a Zeiss LSM 5 PASCAL confocal la-
ser scanning microscope, and in double-labeling experiments, the
multi-tracking mode was selected, unless otherwise noted.
2855
2.5. Interaction of preincubated hIAPP with cells
Ten micromolar hIAPP was preincubated in medium for 24 h at
37 °C with 5% CO2. The incubated peptides were directly applied to
plated PC12 cells and further incubated for 30 min at 37 °C with 5%
CO2. GM1 labeling was performed as described above.
2.6. Inhibition of gangliosides or cholesterol synthesis
PC12 cells were incubated with 20 lM PDMP for 24 h or 1 lM
compactin for 48 h, which inhibited gangliosides or cholesterol
synthesis, respectively. The compactin-treatment was reported to
reduce the cellular cholesterol level to about 40% of the normal le-
vel without affecting the viability [19]. The reagent-containing
medium was replaced with fresh medium containing 20 lM hIAPP
and the cells were incubated for 24 h. Cell staining was performed
as described above.
2.7. Estimation of ganglioside content and accumulated amount of
hIAPP amyloids
The amount of cell surface gangliosides and accumulated hIAPP
amyloids were estimated by the fluorescence intensity of bound
CTX-B and Congo red, respectively [20,21]. The background fluo-
rescence was subtracted from the CTX-B or Congo red fluorescence
expressed in 256 tones pixel by pixel. The intensity multiplied by
the pixel number was summed and normalized to the cell number
examined. This value reflects the quantity of gangliosides or accu-
mulated hIAPP amyloids per cell.
3. Results and discussion
Ganglioside-expressing PC12 cells were incubated with 10 lM
hIAPP for 24 h, and subsequently stained with Congo red and
CTX-B, the former being suitable for staining amyloids in live cells
[13] and the latter being a specific probe for gangliosides. The dis-
tribution of gangliosides on PC12 cell membranes is heterogenous
(Fig. 2A, PC12 untreated). Ganglioside-rich domains, in which gan-
gliosides are brightly stained with CTX-B, are also cholesterol-rich
and abolished by cholesterol depletion [13,22], suggesting that the
microscopic domains were assemblies of nano-scale membrane
patches, called ‘lipid raft’ [23]. hIAPP specifically accumulated in
the ganglioside- and cholesterol-rich domains in PC12 cell mem-
branes, forming amyloid fibrils (Fig. 1A). Fig. 1B shows the concen-
tration-dependent accumulation of the amyloid. At a concentration
of 25 lM, hIAPP was clearly cytotoxic, as judged from differential
interference contrast (DIC) images and fluorescence micrographs
of a live cell indicator, calcein-AM (data not shown). The peptide-
induced cytotoxicity was observed after hIAPP amyloid had accu-
mulated on cell membranes to some extent, suggesting that the
native cytotoxic form of hIAPP was produced during the process
of the peptide aggregation into Congo red-positive amyloids. Addi-
tionally, on the pathway of amyloidogenesis, hIAPP does not form
small oligomers [24], indicating that the cytotoxic form was possi-
bly amyloid fibrils.
To examine whether hIAPP initially bound to and then accumu-
lated on cell membranes, or initially aggregated in the medium and
then bound to the membranes, hIAPP was preincubated in medium
for 24 h and then applied to PC12 cells. No accumulation of amy-
loid was observed (Fig. 1C), indicating that hIAPP formed amyloids
on ganglioside- and cholesterol-rich domains and fibrils formed in
solution were not the main toxic components.
To investigate whether gangliosides and cholesterol were
essential for the accumulation of hIAPP amyloid on cell mem-
branes, PC12 cells were treated with PDMP, an inhibitor of gangli-
2856 M. Wakabayashi, K. Matsuzaki / FEBS Letters 583 (2009) 2854–2858
Fig. 1. Accumulation of hIAPP amyloids on PC12 cell membranes. (A) PC12 cells were incubated with 10 lM hIAPP for 24 h and stained with Congo red and CTX-B. (B) PC12
cells were incubated with hIAPP at the indicated concentrations for 24 h and stained with Congo red. (C) Ten micromolar hIAPP was preincubated in culture medium for 24 h,
and then applied to PC12 cells for 30 min.
oside synthesis, or compactin, an inhibitor of cholesterol synthesis,
and then incubated with 20 lM hIAPP. PDMP significantly reduced
the amount of ganglioside to about 43% of control (Fig. 2A). The
compactin-treatment was reported to reduce the cellular choles-
terol level to about 40% of the normal level without affecting the
viability [19] and we have found that inhibiting the synthesis of
cholesterol reduced ganglioside content [13]. According to the de-
crease in ganglioside and cholesterol contents, the accumulation of
hIAPP amyloids was significantly suppressed (Fig. 2B). PDMP- and
compactin-treatment lowered the amount of hIAPP accumulated
to 28% and 36% of the control level, respectively.
CHO cells, which express no ganglioside, were also incubated
with 20 lM hIAPP for 24 h, and subsequently stained with Congo
red and CTX-B. CHO cell membranes were not stained with CTX-
B (Fig. 2A), and hIAPP amyloid did not accumulate on the mem-
branes (Fig. 2B).
These results clearly showed that gangliosides and cholesterol
played crucial roles in the formation of amyloid fibrils by hIAPP. Both
gangliosides and cholesterol are components of lipid rafts in plasma
membrane and the depletion of each lipid reduced the accumulation
of amyloid, evidently indicating that gangliosides in lipid rafts were
the cause of the formation of amyloid by hIAPP. The ganglioside-in-
duced production of amyloid fibrils is also supported by the findings
that amounts of gangliosides were markedly greater in pancreatic is-
lets than in pancreatic tissues [25] and the deposition of hIAPP amy-
loid was specific to pancreatic islet b cells [8,9].
Sparr et al. demonstrated using rat insulinoma tumor (RIN) cell
that the presence of cell membranes promoted the formation of
amyloid by hIAPP and the amyloidogenesis was coupled to the
extraction of membrane lipids and the subsequent permeabiliza-
tion of the lipid membranes, leading to cytotoxicity [26]. Recently,
the uptake of membrane lipids of pancreatic b-cell line INS-1E dur-
ing the aggregation of hIAPP was also reported [27]. In the case of
Alzheimer’s amyloid b-protein, ganglioside-bound form promotes
the protein aggregation and cytotoxicity [28]. In this study, PC12
cell membranes containing gangliosides induced the amyloid for-
mation by hIAPP.
Moreover, it was reported that the amino acid sequences
responsible for amyloid formation and membrane disruption are
located in different regions of the peptide and amyloid formation
can occur independently from membrane disruption [29,30]. The
C-terminal domain has amyloidogenic property, and the N-termi-
nal fragment of hIAPP was alone sufficient to induce membrane
perturbation and a single mutation abolished the activity [31–
33]. The interaction of the N-terminal part of hIAPP with mem-
branes is driven by electrostatic interactions. However, model lipid
membranes used in these reports include only PG as an anionic li-
pid, which is not expressed in the outer leaflets of plasma mem-
branes of mammalian cells. Another anionic lipid, PS, has also
been proposed as an inducer of amyloid formation and subsequent
cell death by hIAPP and amyloid b-protein [12,34,35]. However, PS
constitutively exists in the inner leaflets and its exposure to the
outer leaflets implies apoptotic cell death, suggesting that the
interaction of the peptides with PS may not be considered as the
primary cause of extracellular amyloid deposits and cell death in
intact islet cells. Therefore, the interaction of N-terminal region
of hIAPP with the membranes would be mediated by gangliosides,
which are major anionic lipids of mammalian cell membranes.
Gangliosides in lipid rafts are clustered [36] and the local electro-
static interaction with hIAPP would be significantly enhanced.
These findings possibly indicate the existence of two independent
processes. hIAPP preferentially binds to gangliosides in lipid rafts
by electrostatic interactions and the ganglioside-bound peptide
facilitates the amyloid formation. During amyloid formation pro-
cess, hIAPP takes up membrane lipids, permeabilizes the mem-
branes, and leads to cell death. Accelerated binding of hIAPP to
cell surface gangliosides ultimately, albeit indirectly, elicits the en-
hanced cytotoxicity. We previously reported that ganglioside-
bound form of amyloid b-protein generates negative curvature
strain on lipid bilayers [37]. hIAPP also provokes negative mem-
 M. Wakabayashi, K. Matsuzaki / FEBS Letters 583 (2009) 2854–2858 2857

Fig. 2. Gangliosides- and cholesterol-dependent accumulation of hIAPP amyloids on plasma membranes. (A) Cell surface gangliosides of PC12 or CHO cells in the indicated
conditions were stained with CTX-B. Ganglioside contents were estimated from CTX-B fluorescence intensity. See Section 2 for details. The data shown are the means ± S.D.
(n = 4). P(untreated versus PDMP-treated) < 0.01. The statistical analysis was performed by the two-sample t-test. (B) PC12 or CHO cells in the indicated conditions were
incubated with 20 lM hIAPP for 24 h and then stained with Congo red. The amount of accumulated hIAPP amyloid was estimated from Congo red fluorescence intensity. See
Section 2 for details. The data shown are the means ± S.D. (n = 3). P(untreated versus PDMP-treated) < 0.02, P(untreated versus compactin-treated) < 0.02. The statistical
analysis was performed by the two-sample t-test.

brane curvature [38], suggesting that the membrane distortion
might contribute to the hIAPP-induced cytotoxic effect.
In conclusion, we demonstrated that gangliosides in lipid rafts
serve as a platform for the formation of amyloid deposits and the
resultant cytotoxic activity not only with Alzheimer’s amyloid b-
protein [13] but also with hIAPP suggesting this raft-mediated
amyloidogenesis to be a universal mechanism for amyloidogenic
proteins.
References
[1] Armstrong, R.A., Cairns, N.J. and Lantos, P.L. (2001) Spatial pattern of prion
protein deposits in patients with sporadic Creutzfeldt–Jakob disease.
Neuropathology 21, 19–24.
[2] Forloni, G. (1996) Neurotoxicity of beta-amyloid and prion peptides. Curr.
Opin. Neurol. 9, 492–500.
[3] Marzban, L., Park, K. and Verchere, C.B. (2003) Islet amyloid polypeptide and
type 2 diabetes. Exp. Gerontol. 38, 347–351.
[4] Selkoe, D.J. (1991) The molecular pathology of Alzheimer’s disease. Neuron 6,
487–498.
[5] Harper, J.D. and Lansbury Jr., P.T. (1997) Models of amyloid seeding in
Alzheimer’s disease and scrapie: mechanistic truths and physiological
consequences of the time-dependent solubility of amyloid proteins. Annu.
Rev. Biochem. 66, 385–407.
[6] Cooper, G.J., Willis, A.C., Clark, A., Turner, R.C., Sim, R.B. and Reid, K.B.
(1987) Purification and characterization of a peptide from amyloid-rich
pancreases of type 2 diabetic patients. Proc. Natl. Acad. Sci. USA 84, 8628–
8632.
[7] Westermark, P., Wernstedt, C., Wilander, E., Hayden, D.W., O’Brien, T.D. and
Johnson, K.H. (1987) Amyloid fibrils in human insulinoma and islets of
Langerhans of the diabetic cat are derived from a neuropeptide-like protein
also present in normal islet cells. Proc. Natl. Acad. Sci. USA 84, 3881–3885.
[8] Kahn, S.E., Andrikopoulos, S. and Verchere, C.B. (1999) Islet amyloid: a long-
recognized but underappreciated pathological feature of type 2 diabetes.
Diabetes 48, 241–253.
[9] Rocken, C., Linke, R.P. and Saeger, W. (1992) Immunohistology of islet amyloid
polypeptide in diabetes mellitus: semi-quantitative studies in a post-mortem
series. Virchows Arch. A Pathol. Anat. Histopathol. 421, 339–344.
[10] Gorbenko, G.P. and Kinnunen, P.K. (2006) The role of lipid–protein interactions
in amyloid-type protein fibril formation. Chem. Phys. Lipids 141, 72–82.
[11] Domanov, Y.A. and Kinnunen, P.K. (2008) Islet amyloid polypeptide forms
rigid lipid–protein amyloid fibrils on supported phospholipid bilayers. J. Mol.
Biol. 376, 42–54.
[12] Engel, M.F. et al. (2008) Membrane damage by human islet amyloid
polypeptide through fibril growth at the membrane. Proc. Natl. Acad. Sci.
USA 105, 6033–6038.
[13] Wakabayashi, M. and Matsuzaki, K. (2007) Formation of amyloids by Abeta-
(1–42) on NGF-differentiated PC12 cells: roles of gangliosides and cholesterol.
J. Mol. Biol. 371, 924–933.
[14] Simons, K. and Ikonen, E. (1997) Functional rafts in cell membranes. Nature
387, 569–572.
[15] Manes, S., del Real, G. and Martinez, A.C. (2003) Pathogens: raft hijackers. Nat.
Rev. Immunol. 3, 557–568.
[16] Hanzal-Bayer, M.F. and Hancock, J.F. (2007) Lipid rafts and membrane traffic.
FEBS Lett. 581, 2098–2104.
[17] Simons, K. and Toomre, D. (2000) Lipid rafts and signal transduction. Nat. Rev.
Mol. Cell Biol. 1, 31–39.
[18] Rhoades, E., Agarwal, J. and Gafni, A. (2000) Aggregation of an amyloidogenic
fragment of human islet amyloid polypeptide. Biochim. Biophys. Acta 1476,
230–238.
[19] Wang, S.S., Rymer, D.L. and Good, T.A. (2001) Reduction in cholesterol and
sialic acid content protects cells from the toxic effects of beta-amyloid
peptides. J. Biol. Chem. 276, 42027–42034.
2858 M. Wakabayashi, K. Matsuzaki / FEBS Letters 583 (2009) 2854–2858
[20] Kiyokawa, E., Baba, T., Otsuka, N., Makino, A., Ohno, S. and Kobayashi, T. (2005)
Spatial and functional heterogeneity of sphingolipid-rich membrane domains.
J. Biol. Chem. 280, 24072–24084.
[21] Nishio, M., Fukumoto, S., Furukawa, K., Ichimura, A., Miyazaki, H., Kusunoki, S.,
Urano, T. and Furukawa, K. (2004) Overexpressed GM1 suppresses nerve
growth factor (NGF) signals by modulating the intracellular localization of
NGF receptors and membrane fluidity in PC12 cells. J. Biol. Chem. 279, 33368–
33378.
[22] Wakabayashi, M., Okada, T., Kozutsumi, Y. and Matsuzaki, K. (2005) GM1
ganglioside-mediated accumulation of amyloid beta-protein on cell
membranes. Biochem. Biophys. Res. Commun. 328, 1019–1023.
[23] Jacobson, K., Mouritsen, O.G. and Anderson, R.G. (2007) Lipid rafts: at a
crossroad between cell biology and physics. Nat. Cell Biol. 9, 7–14.
[24] Soong, R., Brender, J.R., Macdonald, P.M. and Ramamoorthy, A. (2009)
Association of highly compact type II diabetes related islet amyloid
polypeptide intermediate species at physiological temperature revealed by
diffusion NMR spectroscopy. J. Am. Chem. Soc. 131, 7079–7085.
[25] Saito, M. and Sugiyama, K. (2000) A distinct ganglioside composition of rat
pancreatic islets. Arch. Biochem. Biophys. 376, 371–376.
[26] Sparr, E., Engel, M.F., Sakharov, D.V., Sprong, M., Jacobs, J., de Kruijff, B.,
Hoppener, J.W. and Killian, J.A. (2004) Islet amyloid polypeptide-induced
membrane leakage involves uptake of lipids by forming amyloid fibers. FEBS
Lett. 577, 117–120.
[27] Radovan, D., Opitz, N. and Winter, R. (2009) Fluorescence microscopy studies
on islet amyloid polypeptide fibrillation at heterogeneous and cellular
membrane interfaces and its inhibition by resveratrol. FEBS Lett. 583, 1439–
1445.
[28] Okada, T., Wakabayashi, M., Ikeda, K. and Matsuzaki, K. (2007) Formation of
toxic fibrils of Alzheimer’s amyloid beta-protein-(1–40) by
monosialoganglioside GM1, a neuronal membrane component. J. Mol. Biol.
371, 481–489.
[29] Brender, J.R., Lee, E.L., Cavitt, M.A., Gafni, A., Steel, D.G. and Ramamoorthy, A.
(2008) Amyloid fiber formation and membrane disruption are separate
processes localized in two distinct regions of IAPP, the type-2-diabetes-
related peptide. J. Am. Chem. Soc. 130, 6424–6429.
[30] Lopes, D.H., Meister, A., Gohlke, A., Hauser, A., Blume, A. and Winter, R. (2007)
Mechanism of islet amyloid polypeptide fibrillation at lipid interfaces studied
by infrared reflection absorption spectroscopy. Biophys. J. 93, 3132–3141.
[31] Brender, J.R., Hartman, K., Reid, K.R., Kennedy, R.T. and Ramamoorthy, A.
(2008) A single mutation in the nonamyloidogenic region of islet amyloid
polypeptide greatly reduces toxicity. Biochemistry 47, 12680–12688.
[32] Nanga, R.P., Brender, J.R., Xu, J., Hartman, K., Subramanian, V. and
Ramamoorthy, A. (2009) Three-dimensional structure and orientation of rat
islet amyloid polypeptide protein in a membrane environment by solution
NMR spectroscopy. J. Am. Chem. Soc. 131, 8252–8261.
[33] Nanga, R.P., Brender, J.R., Xu, J., Veglia, G. and Ramamoorthy, A. (2008)
Structures of rat and human islet amyloid polypeptide IAPP(1–19) in micelles
by NMR spectroscopy. Biochemistry 47, 12689–12697.
[34] Ciccotosto, G.D., Tew, D.J., Drew, S.C., Smith, D.G., Johanssen, T., Lal, V., Lau, T.L.,
Perez, K., Curtain, C.C., Wade, J.D., Separovic, F., Masters, C.L., Smith, J.P.,
Barnham, K.J. and Cappai, R. (2009). Stereospecific interactions are necessary
for Alzheimer disease amyloid-beta toxicity. Neurobiol. Aging, in press.
[35] Lau, T.L., Gehman, J.D., Wade, J.D., Masters, C.L., Barnham, K.J. and Separovic, F.
(2007) Cholesterol and Clioquinol modulation of A beta(1–42) interaction
with phospholipid bilayers and metals. Biochim. Biophys. Acta 1768, 3135–
3144.
[36] Kakio, A., Nishimoto, S.I., Yanagisawa, K., Kozutsumi, Y. and Matsuzaki, K.
(2001) Cholesterol-dependent formation of GM1 ganglioside-bound amyloid
beta-protein, an endogenous seed for Alzheimer amyloid. J. Biol. Chem. 276,
24985–24990.
[37] Matsuzaki, K. and Horikiri, C. (1999) Interactions of amyloid beta-peptide
(1–40) with ganglioside-containing membranes. Biochemistry 38, 4137–
4142.
[38] Smith, P.E., Brender, J.R. and Ramamoorthy, A. (2009) Induction of negative
curvature as a mechanism of cell toxicity by amyloidogenic peptides: the case
of islet amyloid polypeptide. J. Am. Chem. Soc. 131, 4470–4478.