parkinson s 
 disease is one of several amyloid disorders , 
 collectively referred to as synucleinopathies , whose pathology is 
 characterized by the aggregation of the presynaptic protein -synuclein 
 ( -syn ) into lewy bodies . despite the appearance of these 
 lewy bodies in diseased brains , soluble oligomers of -syn the characterization of -syn oligomers is an outstanding 
 biophysical challenge due to their heterogeneity and propensity to 
 aggregate . these properties have precluded -syn oligomers from 
 high - resolution structural characterization by x - ray crystallography 
 and have limited their characterization to a range of low - resolution 
 techniques , including size - exclusion chromatography , sds - page , dynamic 
 light scattering , analytical ultracentrifugation , and cryo - tem . the same properties have precluded oligomers 
 formed by many amyloidogenic proteins from structural characterization 
 at high resolution . small peptides derived from amyloidogenic 
 proteins have afforded 
 high - resolution structures that provide insights into the structures 
 of amyloid oligomers formed by full - length proteins . studying the assembly of small peptides derived from -syn 
 may provide insights into oligomeric assemblies of the protein . several 
 structural studies of -syn oligomers have suggested that two 
 -strands loosely defined by residues 3643 and 4958 
 form the core of the toxic oligomers associated with parkinson s 
 disease . most notably , hoyer et al . recently observed a -hairpin 
 defined by residues 3655 in monomeric -syn by nmr spectroscopy 
 ( figure 1a ) . the authors found that sequestering this -hairpin 
 in an engineered binding protein markedly reduces the toxicity of 
 aged -syn and inhibits its fibrillization . five of the six known disease - causing point mutations of parkinson s 
 disease are located within this -hairpin , further emphasizing 
 the importance that this region plays in the pathology of parkinson s 
 disease . recently , schulten et al . have found through molecular dynamics 
 simulations that residues 3655 adopt a -hairpin similar 
 to that observed by hoyer et al . the 
 authors also found that disease - causing point mutations stabilize 
 the -hairpin . they further suggest that -hairpin formation 
 precedes aggregation of -syn in the pathway to pathology . the 
 concurrence of genetic and structural evidence motivated us to design 
 a macrocyclic -sheet that mimics this -hairpin , with 
 the goal of creating a high - resolution structural model of -syn 
 oligomers ( figure 1 ) . ( a ) nmr structure of the -hairpin 
 formed by residues 3655 in full - length -syn ( green ) 
 bound by an engineered affibody ( white ) ( pdb 4bxl ) . we designed macrocyclic 
 -sheet peptide 1a to 
 mimic the -hairpin formed by -syn3655 ( figure 1b and c ) : 
 we incorporated the heptapeptides -syn3642 ( gvlyvgs ) and -syn4955 ( vhgvatv ) into 
 the top and bottom strands of the macrocycle to maintain the same 
 alignment and hydrogen - bonding patterns observed in the nmr structure . we replaced the residues that form the loop of the -hairpin 
 ( 4348 ) with a -linked ornithine turn unit , which serves 
 as a -turn mimic and enforces a -sheet conformation . we connected residues 36 and 55 with a second 
 -linked ornithine turn to further enforce a -sheet conformation . we incorporated a single n - methyl group 
 on val52 to limit the uncontrolled aggregation of peptide 1a . we mutated tyr39 to 4-iodophenylalanine ( phe ) to allow x - ray crystallographic 
 phase determination using single wavelength anomalous dispersion ( sad ) 
 phasing . this approach has allowed us to determine the x - ray 
 crystallographic 
 structure of oligomers formed by this -hairpin derived from 
 -syn3655 . in 
 this structure , we observe a hierarchical assembly of -hairpins : 
 three -hairpins assemble to form a trimer , three trimers assemble 
 to form a nonamer , and two nonamers pack to form an octadecamer . this 
 structure is the first reported x - ray crystallographic structure of 
 oligomeric assemblies of peptides derived from -syn . these 
 oligomers suggest a model for -syn oligomerization in which 
 self - assembly of -syn is centered around -syn3655 . peptide 1a and its derivatives 
 were synthesized using fmoc - based solid - phase peptide synthesis ( scheme s1 ) . screening 
 peptide 1a in 288 conditions yielded a single condition 
 in which crystals grew : 0.1 m hepes buffer at ph 8.0 , 0.5 m ( nh4)2so4 , and 34% 2-methyl-2,4-pentanediol 
 ( mpd ) . diffraction data were collected to 1.97  at the advanced 
 light source at lawrence berkeley national laboratory with a synchrotron 
 source at 0.976  . the locations of the anomalous scattering atoms 
 were determined using the program hyss ( hybrid structure search ) . the structure of peptide 1a was 
 solved and refined in space group p213 . the asymmetric unit contains six distinct 
 copies of peptide 1a , each of which is folded into an 
 antiparallel -hairpin with minor differences in the conformations 
 of thr54 , val55 , and the -linked ornithine 
 turn unit that joins ala36 to val55 ( figures 2 , s1 , and s2 ) . the residues of the -hairpin are displayed 
 on the front or back surfaces of the -sheet : ala36 , leu38 , val40 , ser42 , val49 , gly51 , ala53 , and val55 are displayed 
 on the front surface ; val37 , phe39 , gly41 , his50 , n - me val52 , and thr54 are displayed on the back surface . the -hairpin monomers have a right - handed 
 twist ranging from approximately 1530 per residue along 
 the -strand axis , thus mimicking the highly twisted -hairpin 
 observed by hoyer et al . ( a ) representative -hairpin monomer from 
 the crystal lattice of peptide 1a ( pdb 5f1 t ) . ( b ) -hairpin 
 monomer formed by -syn3655 ( pdb 4bxl , affibody omitted ) . the differences in conformations of thr54 appear 
 to 
 be responsible for the slight differences among the -hairpin 
 monomers . four of the six monomers in the asymmetric unit form an 
 ideal -hairpin . in the fifth monomer , the hydroxy group of 
 thr54 is positioned such that it disrupts 
 the interchain hydrogen bond between the amide proton of val55 and the carbonyl oxygen of ala36 ( figure s2b ) . in the sixth monomer , both of these conformations abrogate 
 the intramolecular hydrogen bonding between ala36 and val55 and distort the conformation of the -linked ornithine 
 turn that connects them . the six -hairpin monomers of 
 the asymmetric unit further 
 assemble into two triangular trimers in which three monomers occupy 
 the edges of the triangle ( figure 3a ) . each 
 trimer is composed of two monomers in one orientation and one monomer 
 in a different orientation . the relative orientations of the n - methyl groups within the trimer highlight the lack of 
 internal symmetry ; two of the n - methyl groups point 
 into the center of the trimer , whereas the third points outward ( figure 3b ) . ( a ) cartoon and stick 
 representation depicting the intramolecular and intermolecular hydrogen 
 bonding within the trimer ( front surface view ) . ( b ) cartoon representation 
 depicting the location of the n - methyl groups in 
 the trimer ( front surface view ) . ( c ) sphere representation depicting 
 the hydrophobic packing of residues in the trimer ( front surface view ) . . 1300  of surface area per 
 trimer , averaging 450  of buried surface area per 
 -hairpin monomer ( figure 3c ) . this area corresponds to 
 more than one - fourth of the total surface area of the monomers . the 
 two surfaces of each trimer display different residues : the front 
 surface of each trimer displays the residues on the front surface 
 of the -hairpin monomers ; the back surface of each trimer displays 
 the residues on the back surface -hairpin monomers . intermolecular 
 hydrogen bonds at the vertices of the trimer further stabilize this 
 assembly . the two crystallographically distinct trimers further 
 assemble 
 to form two distinct basket - shaped nonamers , each of which is a trimer 
 of the triangular trimers . unlike 
 the trimer subunit , each nonamer contains internal c3 symmetry , resulting in uniform packing of the trimer subunits against 
 one another . hydrogen bonding between the trimer subunits stabilizes 
 the basket - shaped nonamer ( figure 4b ) . the vertices of the trimer subunits within the 
 nonamers form extensive networks of hydrogen bonds . at the juncture 
 of each of the trimers , four -hairpins come together to form 
 an eight - stranded -barrel ( figure s3 ) . the top of the 
 basket - shaped nonamer comprises a hydrogen - bonded triangular interface 
 ( figure 4b ) . in this 
 interface , each triangular trimer subunit contributes one -hairpin 
 and each of these -hairpins forms six intermolecular hydrogen 
 bonds . ( b ) cartoon and stick 
 representation depicting the main - chain hydrogen - bonding networks 
 ( outer surface view ) . ( c ) hydrophobic packing in the core of the nonamer 
 ( inner surface view , image is rotated 180 with respect to a 
 and b about the vertical axis ) . hydrophobic contacts between the triangular trimers also 
 stabilize 
 each nonamer ( figure 4c ) . the trimer subunits pack against each other along their back 
 surfaces to form a densely packed hydrophobic core consisting of residues 
 val37 , phe39 , his50 , 
 val52 , and thr54 . 3000  of 
 surface area in the nonamer assembly : nearly 1000  per trimer subunit . this area corresponds to roughly one - third of 
 the total surface area of the trimer . the nonamers 
 pack against one another through hydrophobic contacts between ala36 , leu38 , val40 , val55 , and 
 the -linked ornithine turn unit that connects ala36 to val55 ( figure 5b ) . . 1300  of surface 
 area , of which each nonamer contributes roughly 650  . contacts between octadecamers within the lattice are small , 
 roughly 300  per octadecamer . inspired by the interest in racemic and enantiomeric proteins , 
 we also determined the x - ray crystallographic structure of peptide ent-1a . as expected , peptide ent-1a crystallizes from the same conditions 
 as peptide 1a and forms crystals in the same space group 
 with similar unit cell dimensions as those formed by peptide 1a . the asymmetric unit of peptide ent-1a contains six molecules of ent-1a , each of which is folded into a -hairpin that is the mirror 
 image of that formed by peptide 1a ( figure 6 ) . peptide ent-1a assembles to form oligomers that are mirror images 
 of those formed by peptide 1a . these enantiomeric oligomers 
 are identical in every fashion , except handedness , to those formed 
 by peptide 1a . although racemates often crystallize more 
 readily than individual enantiomers , attempts to cocrystallize peptides 1a and ent-1a have failed . monomers from the crystal lattice of peptide ent-1a ( left , pdb 5f1w ) and peptide 1a ( right , pdb 5f1 t ) . we envisioned that full - length -syn could 
 assemble in the same fashion as the oligomers formed by peptide 1a , but were concerned whether the trimers and nonamers would 
 accommodate the loop and additional n- and c - terminal residues . to address this question we built 
 residues 4348 ( ktkegv ) , 3335 ( tke ) , and 5658 
 ( aek ) into the crystallographic coordinates of peptide 1a and performed replica - exchange molecular dynamics ( remd ) to generate 
 realistic conformations of the loops and the n- and c - terminal fragments of the -hairpin . the remd simulation shows that the nonamer successfully accommodates 
 the additional residues from the full - length protein without any significant 
 clashes among residues ( figure 7 ) . the n- and c - terminal fragments of -syn project 
 out of the assembly and do not interfere with nonamer formation . the 
 residues on each of the loops at the vertices of the nonamer pack 
 against one another , suggesting that these additional residues from 
 the full - length protein could stabilize this assembly . incorporation 
 of the loops as well as the additional n- and c - terminal residues 
 into the nonamer buries an additional 400  of surface 
 area per trimer subunit , providing 
 an additional 1200  of buried surface area beyond 
 the crystallographic nonamer . table 1 summarizes the size of the contact surfaces within 
 the crystallographic oligomers and the model of -syn3358 . the x - ray crystallographic and remd structures generated herein 
 may serve as models for the core of the oligomers formed by full - length 
 -syn . the interaction of 
 -syn with the 
 anionic membranes of neurons induces conformational changes and nucleates 
 self - assembly of the protein . to test 
 the effects of membranes on the conformation of peptide 1a , we compared the circular dichroism ( cd ) spectra of peptide 1a in the presence of anionic or neutral liposomes to that 
 of peptide 1a in aqueous buffer ( figure 8) . the cd spectrum of peptide 1a in aqueous buffer displays negative bands centered at 220 
 and 200 nm . upon the addition of anionic 
 phosphatidylcholine : phosphatidylserine ( pc : ps ) large unilamellar vesicles 
 ( luvs ) , the cd spectrum of peptide 1a changes dramatically : 
 the negative band at 220 nm becomes more intense and a positive band 
 appears below ca . this change in cd spectrum upon addition 
 of the liposomes indicates that peptide 1a adopts a -sheet - rich 
 conformation upon interaction with anionic pc : ps liposomes . spectra of 50 m 
 peptide in 10 mm sodium phosphate buffer at ph 7.4 were acquired in 
 the presence or absence of 1.0 mm lipids , constituting either phosphatidylcholine 
 ( pc ) or phosphatidylcholine : phosphatidylserine ( pc : ps ) liposomes . in contrast to anionic pc : ps luvs , 
 neutral phosphatidylcholine 
 ( pc ) luvs do not induce changes in the conformation of peptide 1a . the cd spectrum of peptide 1a in the presence 
 of pc luvs is superimposable with the cd spectrum of peptide 1a in aqueous buffer . the contrasting 
 effects of the pc and pc : ps liposomes indicates that electrostatic 
 interactions between the cationic peptide and the anionic liposomes 
 are essential to the membrane - induced conformational changes observed . to test the importance of chirality in the interaction of peptide 1a with pc : ps liposomes , we investigated the effect of liposomes 
 on the cd spectra of peptide ent-1a . the cd spectra of peptide ent-1a are 
 identical but opposite in sign to those of peptide 1a in the three sets of conditions studied . in aqueous buffer , the 
 cd spectrum of peptide ent-1a displays 
 positive bands centered at 220 and 200 nm . in the presence of pc : ps 
 luvs , the positive band at 220 nm becomes more intense and a negative 
 band appears below ca . no change in the cd spectrum is observed 
 upon the addition of pc luvs . the equivalent behavior of the enantiomeric 
 peptide suggests that chiral interactions are not important in the 
 interaction with liposomes , even though the individual lipid molecules 
 are chiral . instead , the interaction appears to reflect the importance 
 of the charged head groups and the hydrophobic lipids . the oligomers formed by full - length -syn are thought 
 to induce cell death upon interaction with the cell membrane . to determine whether the propensity of peptide 1a to bind lipid membranes imparts toxicity , we studied the 
 effect of peptide 1a and several control peptides on 
 a neuronally derived cell line . treatment of sh - sy5y cells with peptide 1a results in cell death as measured by lactate dehydrogenase 
 release ( ldh ) and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium 
 bromide ( mtt ) conversion assays ( figure 9 ) . a dose - response relationship is observed 
 at concentrations ranging from 540 m , with maximal 
 toxicity occurring at 40 m . maximal cell death occurs within 
 16 h after treatment with 40 m peptide 1a ( figure s4 ) . these results 
 demonstrate that peptide 1a is cytotoxic but do not establish 
 that the toxicity results from membrane interaction . ldh and mtt assays of 
 the toxicity of peptide 1a toward 
 sh - sy5y cells . cells were incubated with 540 m of peptide 1a for 24 h before performing the assays . to establish whether cell death 
 occurs from membrane interaction , 
 we compared peptide 1a to three homologues ( ent-1a , 1aa53e , and 1b ) in ldh and mtt assays ( figure 10 ) . treatment of sh - sy5y cells with either 
 peptide ent-1a or peptide 1a results in nearly equal levels of cell death . the comparable toxicity 
 of peptides ent-1a and 1a suggest that toxicity results from nonspecific interactions with 
 the cells , rather than through specific interaction with a receptor 
 protein . ldh and mtt assays of the toxicity of peptide 1a and 
 its homologues toward sh - sy5y cells . cells were incubated with 40 
 m of peptide 1a , ent-1a , 1aa53e , or 1b for 24 h before performing the assays . cell death and viability 
 were determined spectrophotometrically as previously described . peptide 1aa53e is identical 
 to peptide 1a in amino acid sequence with exception of 
 the a53e familial point mutation . this mutation reduces the affinity 
 of full - length -syn to anionic lipid membranes and delays its 
 aggregation . cd spectroscopy shows that 
 this mutation blocks interactions between peptide 1aa53e and anionic lipid membranes : the cd spectra 
 of peptide 1aa53e are identical 
 in the presence or absence of pc : ps luvs ( figure s5 ) . peptide 1aa53e produces 
 no measurable cell death at 40 um ( figure 10 ) . the lack of toxicity of peptide 1aa53e in conjunction with its lack 
 of membrane interactions supports that membrane interaction is central 
 to the toxicity of peptide 1a . peptide 1b was designed 
 to test the role of the crystallographically 
 observed oligomers in the toxicity elicited by peptide 1a . peptide 1b is identical to peptide 1a in amino acid sequence , but is n - methylated at 
 gly41 instead of val52 . we anticipated that n - methylation at gly41 would disrupt the formation 
 of the trimer and the basket - shaped nonamer by disrupting the hydrogen - bonding 
 within the oligomers and forcing the subunits apart ( figure s6 ) . peptide 1b is much less cytotoxic 
 than peptide 1a , as determined by both the ldh and mtt 
 assays ( figure 10 ) . n - methylation at gly41 does not impair the interaction 
 of peptide 1b and lipid bilayer membranes : upon the addition 
 of pc : ps luvs , the cd spectrum of peptide 1b changes 
 dramatically , in a fashion similar to the changes observed for peptide 1a ( figure s5 ) . the marked decrease 
 in toxicity of peptide 1b is consistent with , but does 
 not prove , the involvement of the crystallographically observed oligomers 
 in the cytotoxicity associated with peptide 1a . the x - ray crystallographic studies of peptide 1a and 
 the molecular modeling studies of the -syn3358 nonamer suggest a model for oligomer formation by full - length -syn . 
 in this model , residues 3655 of full - length -syn form 
 a -hairpin , three of these -hairpins assemble to form 
 triangular trimers , and the trimers assemble to form basket - shaped 
 nonamers or related higher - order oligomers . to our knowledge , the 
 x - ray crystallographic structure of peptide 1a provides 
 the first high - resolution structures of oligomers of peptides derived 
 from -syn . there are currently no reported high - resolution 
 structures of oligomers of the full - length protein . the structures 
 described herein should help bridge the gap between the plethora of 
 low - resolution structural information on full - length -syn oligomers 
 and a still needed high - resolution structure . the structures 
 of the oligomers formed by peptide 1a are fundamentally 
 different from the proposed structure of the fibrils 
 formed by -syn ( figure 11 ) . although there are no high - resolution 
 structures of the fibrils formed by full - length -syn , the current 
 understanding of fibril structures suggests that the monomer units 
 fold onto themselves to form a five - layer -sandwich . in the 
 fibril , the folded monomers hydrogen bond with neighboring monomers 
 to form a network of parallel -sheets . although many of the 
 details of the fibril assembly are still being debated , commonalities 
 such as multilayered parallel -sheets are widely accepted . in contrast to the proposed assembly of the fibril , the oligomers 
 we observe assemble in a different fashion : through the intermolecular 
 interactions among antiparallel -hairpins . this difference 
 may suggest that the oligomers formed by full - length -syn differ 
 from the fibrils in both monomer conformation and in themes of higher - order 
 assembly . ( b ) cartoon of -syn fibril , 
 showing two monomer subunits arranged into a five - layered -sheet . ( c ) cartoon of -syn trimer subunit of 
 basket - shaped nonamers or related higher - order oligomers , showing 
 three -hairpins arranged into a trimer . the x - ray crystallographic structure of peptide 1a suggests an alternative model for oligomer formation by full - length 
 -syn centered around residues 3655 , rather than the 
 nac ( figure 11 ) . residues 
 6195 of -syn , termed the nac , are required for fibrillization of the protein . the currently accepted model 
 of the -syn fibril places the nac at the core of the assembly 
 and -syn3655 at the periphery . the alternative oligomer model places -syn3655 at the core of the oligomers , while the nac and 
 remainder of the protein decorates the periphery . this model does 
 not rule out the possibility of other types of -syn oligomers 
 with the nac at their core . as -syn oligomers are known to 
 be polymorphic , there may be multiple families of oligomers in which 
 different regions of -syn drive assembly . the occurrence 
 of five of the six known familial point mutations 
 in -syn3655 suggests that this region is 
 important in the pathology of synucleinopathies . with exception of 
 h50q , all of these point mutations would be displayed on the solvent - exposed 
 surface of the basket - shaped nonamer . these mutations might stabilize 
 the nonamers or component trimers , or might drive the protein into 
 oligomers with different structures . alternatively , these mutations 
 may merely destabilize native tetramers of -syn and thus promote 
 aggregation . an x - ray crystallographic structure 
 of a derivative of peptide 1a bearing any of these point 
 mutations would be edifying . our own attempts to crystallize homologues 
 of peptide 1a with point mutations h50q , g51d , a53e , 
 or a53 t have thus far been unsuccessful . the oligomers formed 
 by full - length -syn may differ from 
 those formed by peptide 1a while still retaining the 
 general features of -hairpins and trimers . we have , for example , 
 observed that -hairpins derived from -amyloid and 2-microglobulin form symmetrical trimers , rather than the asymmetric 
 trimer observed for peptide 1a . these trimers further assemble to form hexamers , octamers , and dodecamers 
 instead of nonamers and octadecamers . although all of these amyloid - derived 
 peptides differ in sequence and the oligomers differ in precise structure , 
 they share the common theme of -hairpins assembling into triangular 
 trimers that further assemble into complex and intricate higher - order 
 assemblies . trimers may be a unifying motif of toxic amyloid 
 oligomers formed 
 by full - length amyloidogenic proteins . sds - stable trimeric assemblies 
 of full - length -syn have been observed in immunoblots of mouse 
 brain extracts . the appearance of these 
 species strongly correlates with an increased loss of dopaminergic 
 neurons in vivo . similarly , trimeric assemblies of -amyloid 
 have been shown to disrupt cognitive function in rats . the occurrence of the triangular trimeric motif 
 in crystal structures reported by our group has led us to believe 
 that trimers may be unifying substructures of amyloid oligomers that 
 are composed of -hairpin monomers . the cd studies , in conjunction 
 with the toxicity assays , suggest that the cytotoxicity of peptide 1a may result from interactions of trimers or higher - order 
 oligomers with cell membranes , in the same fashion as the oligomers 
 of full - length -syn . at this point , 
 we can not conclusively say that the crystallographic oligomers are 
 forming under the conditions used in the cell assays ; however , the substantial decrease in toxicity of the alternatively n - methylated peptide 1b supports the involvement 
 of the trimer and/or nonamers in cell death . here 
 we present the first x - ray crystallographic structure of oligomers 
 formed by a peptide derived from -syn . this peptide was designed 
 to mimic a -hairpin motif thought to be important in -syn 
 oligomer formation . this designed peptide mimics key properties of 
 oligomers of full - length -syn : affinity for membranes that 
 imparts toxicity . the structure suggests a model for -syn oligomer 
 formation centered around -syn3655 , in which 
 -syn3655 folds into a -hairpin that 
 further assembles into trimers and higher - order oligomers . these structures 
 differ in topology from those of -syn fibrils and suggest that 
 regions of -syn not important for its fibrillization may play 
 a central role in its oligomerization . the x - ray crystallographic 
 structure of peptide 1a and the model of the -syn3358 nonamer may be used as starting points to design 
 small molecules that interact with -syn oligomers or to rationalize 
 the modes of interactions between -syn oligomers and small 
 molecules that interact with them .
oligomeric 
 assemblies of the protein -synuclein are thought 
 to cause neurodegeneration in parkinson s disease and related 
 synucleinopathies . 
 characterization of -synuclein oligomers 
 at high resolution is an outstanding challenge in the field of structural 
 biology . 
 the absence of high - resolution structures of oligomers formed 
 by -synuclein impedes understanding the synucleinopathies at 
 the molecular level . 
 this paper reports the x - ray crystallographic 
 structure of oligomers formed by a peptide derived from residues 3655 
 of -synuclein . 
 the peptide 1a adopts a -hairpin 
 structure , which assembles in a hierarchical fashion . 
 three -hairpins 
 assemble to form a triangular trimer . 
 three copies of the triangular 
 trimer assemble to form a basket - shaped nonamer . 
 two nonamers pack 
 to form an octadecamer . 
 molecular modeling suggests that full - length 
 -synuclein may also be able to assemble in this fashion . 
 circular 
 dichroism spectroscopy demonstrates that peptide 1a interacts 
 with anionic lipid bilayer membranes , like oligomers of full - length 
 -synuclein . 
 ldh and mtt assays demonstrate that peptide 1a is toxic toward sh - sy5y cells . 
 comparison of peptide 1a to homologues suggests that this toxicity results from 
 nonspecific interactions with the cell membrane . 
 the oligomers formed 
 by peptide 1a are fundamentally different than the proposed 
 models of the fibrils formed by -synuclein and suggest that 
 -syn3655 , rather than the nac , may nucleate 
 oligomer formation .