MINIREVIEW
Unraveling the Mechanism of the Vesicle Transport ATPase NSF, the N-Ethylmaleimide-sensitive Factor*

Andrew P. MayDagger §, Sidney W. Whiteheart||, and William I. WeisDagger **

From the Dagger  Departments of Structural Biology and Molecular and Cellular Physiology, Stanford University School of Medicine, Stanford, California 94305 and the  Department of Molecular and Cellular Biochemistry, Chandler Medical Center, University of Kentucky College of Medicine, Lexington, Kentucky 40536

    INTRODUCTION
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INTRODUCTION
Structural Studies
Conformational Responses to...
Mechanisms of SNARE Complex...
Concluding Remarks
REFERENCES

The transport of cargo in eukaryotic cells is mediated by the movement of membranous vesicles that pinch off from one membrane and fuse with another. An essential part of this process is the interaction between SNARE1 (soluble NSF attachment protein receptors) proteins from the vesicle (v-SNARE) and target (t-SNARE) membranes. The resulting SNARE complexes are parallel four-helix coiled-coil structures with melting temperatures between 70 and 90 °C, and it is likely that, at least in part, the free energy of SNARE complex formation drives bilayer fusion (1). Regulating the assembly and disassembly of SNARE complexes is thus an important aspect of vesicular transport.

The hexameric ATPase N-ethylmaleimide-sensitive factor (NSF) uses energy from ATP hydrolysis to dissociate SNARE complexes after membrane fusion, allowing the individual SNARE proteins to be recycled for subsequent rounds of fusion (1). NSF binds to and dissociates SNARE complexes only in the presence of the adaptor protein, alpha -SNAP (soluble NSF attachment protein). alpha -SNAP interacts directly with the SNARE complex and with ATP-bound NSF to form the so-called "20 S particle" (2, 3). In the 20 S particle, alpha -SNAP stimulates the ATPase activity of NSF, leading to SNARE complex disassembly (Fig. 1) (4, 5). Specific v- and t-SNAREs are associated with each intercompartmental transport step, but NSF and alpha -SNAP are general cytosolic factors that can disassemble the SNARE complexes from most, if not all, intracellular transport steps (1).


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Fig. 1.   Schematic representation of the role of NSF in vesicle transport. Vesicles dock and fuse with the target membrane, releasing their cargo. alpha -SNAP and NSF bind to SNARE complexes to form the 20 S complex. Upon hydrolysis of ATP, the individual components of the SNARE complex are released.

The NSF protomer contains three domains: an N-terminal domain, NSF-N (residues 1-205), responsible for interaction with the alpha -SNAP-SNARE complex and two homologous ATP-binding domains, NSF-D1 (residues 206-488) and NSF-D2 (residues 489-744) (3). NSF-D1 is an active ATPase that provides the driving force for SNARE complex disassembly (6, 7). NSF-D1 must bind ATP to interact with the alpha -SNAP-SNARE complex. NSF-D2 is responsible for maintaining NSF as a hexamer (6). It has higher affinity for ATP than NSF-D1 (8) but has no significant ATPase activity. Nucleotide binding by NSF-D2 is, however, important for hexamerization.

The sequences of NSF-D1 and NSF-D2 place NSF in the AAA (ATPases associated with various cellular activities (9)) superfamily (10). AAA proteins, which contain at least one copy of a conserved ~230-amino acid cassette, are involved in a wide variety of cellular roles, including membrane fusion, proteosome regulation, transcription, organelle biogenesis, and microtubule transport and regulation (10, 11). Despite this functional diversity, the ability to assemble or disassemble multisubunit macromolecular complexes, or to fold or unfold polypeptides, appears to be common to the family.

Here, we focus on recent advances that are helping provide a basis for understanding the physical mechanisms that underlie the role of NSF in SNARE complex disruption.

    Structural Studies
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INTRODUCTION
Structural Studies
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Mechanisms of SNARE Complex...
Concluding Remarks
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Crystallographic and electron microscopy studies of NSF, the 20 S complex, as well as other AAA proteins have begun to provide insight into how NSF interacts with the other components of the 20 S complex and responds to changes in bound nucleotide. Electron micrographs of NSF reveal a double ring hexameric structure ~15 nm in diameter and ~12 nm in height (12). The double ring structure is retained by a construct (NSF-D1D2) lacking the NSF-N domain, but the ring height and diameter of NSF-D1D2 are ~2 nm smaller than that of full-length NSF. Similar quaternary arrangements are seen in other AAA proteins (12).

The 20 S complex and alpha -SNAP-SNARE complexes have also been imaged in the electron microscope (12, 13). alpha -SNAP appears to form a sleeve around the SNARE complex, binding lengthwise along the rod-like SNARE coiled-coil (13). The 20 S complex resembles a spark plug, with NSF-D1 and NSF-D2 visible as two rings at one end (Fig. 2A). The alpha -SNAP-SNARE complex sits on the face of the NSF-D1 ring opposite the side that faces NSF-D2. The NSF-D1 hexamer appears flatter and wider than NSF-D2. alpha -SNAP and the SNARE complex interact in an antiparallel manner. This places the membrane-distal, N terminus of the SNARE complex close to the C terminus of alpha -SNAP, which then contacts the N-terminal region of NSF (12, 13).


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Fig. 2.   A, negative stain image of the 20 S complex adapted from Ref. 13. Ribbon diagrams of the components of the 20 S complex are shown approximately to scale. The crystal structure of NSF-D1 is not known, and it is modeled by the structure of NSF-D2. NSF-N domains are shown as the trimer observed in Ref. 14. Only three NSF-N domains are shown for simplicity. Only one copy of alpha -SNAP, based on the structure of Sec17p, is shown for simplicity (there may be up to three copies present in the 20 S complex) (41). The SNARE complex is represented by the structure of a minimal, proteolytically defined complex (34). B, ribbon diagram depicting the structure of the NSF-N domain. The N-terminal subdomain is labeled NN and colored light gray. The C-terminal subdomain is labeled NC and colored dark gray. N- and C termini are labeled. C, top and side views of the p97-ND1 hexamer (17). The subdomains of p97-N and p97-D1 are shown in different colors (p97-N N-terminal subdomain (blue), p97-N C-terminal subdomain (gray), p97-D1 N-terminal subdomain (brown), p97-D1 C-terminal subdomain (khaki)). ADP is shown as white space-filling atoms. The only access to the site equivalent to the putative alpha -SNAP binding site in NSF-N is indicated. D, ribbon diagram of the p97-ND1 monomer. Subdomains are colored as in C. Regions involved in nucleotide binding and potentially transmitting conformational change (N-D1 linker (pink), P-loop (black), DEXX motif (green), sensor-1 (red), sensor-2 (dark brown)) are highlighted. This figure was drawn using the programs BOBSCRIPT (42) and RASTER3D (43).

Crystal structures have been determined for NSF-N (14, 15) and its yeast equivalent Sec18-N (16). The structures of the corresponding domain in p97 (p97-N), a homologue of NSF that functions in NSF-independent membrane trafficking steps (17), and VAT (VAT-N), an archael homologue of p97, are also known (18). NSF-N and its homologues contain two subdomains: an N-terminal beta -barrel and a C-terminal alpha /beta roll. There are significant differences in the structures of the intersubdomain linkers in NSF-N, Sec18-N, p97-N, and VAT-N, but the relative orientations of the two subdomains are virtually identical (14-18). This, together with the demonstration that NSF-N melts with a single sigmoidal transition (15), makes it likely that the two subdomains remain locked together to form a single structural unit.

NSF binds to the alpha -SNAP-SNARE complex only in the presence of ATP. When Sec18p is pretreated with the nonhydrolyzable ATP analogue AMP-PNP, it binds to a cation exchange column, whereas under conditions in which hydrolysis may take place, it does not (16). This suggests that a basic surface is exposed during the conformational change(s) that occur upon ATP binding. The groove formed by the intersubdomain interface in NSF-N is rich in basic and hydrophobic residues and is of appropriate size and shape to accommodate an alpha -helical peptide (15). This region was suggested as a potential site for interaction with the highly negatively charged C-terminal helix of alpha -SNAP (Fig. 2B) (15, 19). Consistent with this, mutation of a conserved arginine residue (Arg67) near the groove ablates NSF binding to alpha -SNAP-SNARE complexes while retaining full ATPase activity.2 Furthermore, hydrophobic residues near the base of the groove have been proposed to interact (16) with a leucine residue required for stimulation of NSF ATPase activity (19), present at the C terminus of alpha -SNAP.

The crystal structure of NSF-D2 (20, 21) consists of an N-terminal nucleotide-binding subdomain and a C-terminal helical subdomain. The NSF-D2 hexamer is formed by packing of the wedge-shaped nucleotide-binding subdomains, with the nucleotide-binding sites located in the interface between protomers. The C-terminal helical subdomains are located at the apices of the hexamer. The structures of both subdomains show similarities to the delta ' clamp-loading subunit of Escherichia coli DNA polymerase III (poldelta ') (22), the protease-associated AAA chaperone, HslU (23, 24), and the D1 ATP-binding domain of p97/VCP (p97-D1) (17). Like NSF-D2, both p97-D1 and HslU form hexamers.

The structure of a fragment of p97 containing the p97-N and p97-D1 domains (p97-ND1) shows that the p97-N domains are located at the periphery of the p97-D1 hexamer (Fig. 2C) (17). Contacts between p97-N and p97-D1 are made with both subdomains of p97-D1 and the N-D1 interdomain linker (Fig. 2D) (17). The relative orientation of the p97-N and p97-D1 domains agrees well with that observed in EM images of VAT (25) and p97 (17, 26). Curiously this orientation looks more like EM images of the ATPgamma S/ATP- than the ADP-bound state of NSF (12), but ADP is bound in the p97-ND1 crystal structure. However, in the p97-ND1 structure, the cleft between the subdomains in p97-N, which includes the surface equivalent to the putative alpha -SNAP binding surface in NSF-N, is blocked by the ND1 linker on the top side of the domain. If this arrangement were true for NSF, the binding cleft would only be accessible on the side opposite the alpha -SNAP-SNARE complex observed in images of the 20 S complex (Fig. 2, A and C). This orientation would not be compatible with binding of the alpha -SNAP-SNARE complex and may explain why the ADP state of NSF is unable to bind alpha -SNAP-SNARE complexes (8).

EM images suggest that the NSF-N domains move with respect to NSF-D1 (12). A conserved glycine residue, present at the C terminus of the N-D1 linker in NSF (and related AAA proteins), may act as a pivot point for rigid body movements between the domains (17). Structurally this glycine residue is in close proximity to the nucleotide-binding site in both NSF-D2 and p97-D1 and could be sensitive to the state of the bound nucleotide. Another glycine residue, found near the N terminus of the p97 N-D1 linker, may contribute to hinge movement (17). However, this residue is not conserved, even among p97 orthologues, and the structures of NSF-N and p97-N diverge in this region.3

    Conformational Responses to Nucleotide State
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Concluding Remarks
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NSF is capable of undergoing large conformational changes upon ATP binding and hydrolysis, most notably with NSF-N domains changing their dispositions relative to the rest of the NSF hexamer (12). In doing so, it converts the energy stored in ATP into mechanical work needed to disassemble the alpha -SNAP-SNARE complex. NSF is a hexamer in solution, and oligomerization appears to be intimately linked with both ATPase activity and ligand binding (3, 7, 11). A construct of NSF containing only the NSF-N and NSF-D1 domains (NSF-ND1) is monomeric, has reduced ATPase activity and affinity for ligand when compared with full-length NSF (6), and is unable to dissociate SNARE complexes. However, when attached to NSF-D2, NSF-D1 forms a hexameric ring, and NSF-D1D2 has ATPase activity comparable with full-length NSF (6). Like NSF, some other active AAA ATPase modules are linked to an oligomerization domain (e.g. FtsH) without which activity is either significantly impaired or lost completely, whereas others (e.g. Vps4p, katanin) exist in equilibrium between monomer and oligomer (11). The stimulation of ATPase activity upon ligand binding and interactions with ligand only when bound to ATP are also common in the AAA family (reviewed in Ref. 11). Oligomerization, ligand binding, conformational change, and ATP hydrolysis therefore appear to cooperate to convert ATP-bound energy to useful work in NSF and other AAA proteins. How are these properties coupled to one another? Although there remains a paucity of direct information on NSF itself, lessons can be learned from recent studies of other members of the AAA family.

AAA proteins contain phosphate-binding P-loop (Walker A) and metal ion-binding DEXX box (Walker B) nucleotide-binding sequences. AAA proteins contain two additional conserved regions, termed sensor-1 and sensor-2 (10, 22), that lie near the nucleotide-binding site. Sensor-1 and sensor-2 are positioned to detect a change in the nucleotide state and to transduce that signal to more remote regions of the protein (10, 20, 22). The structures of two AAA domains that contain consensus sensor-1 (p97-D1 (17)) and sensor-2 (HslU (23, 24)) sequences now provide direct structural information for the sensor regions. However, it remains unclear what role nucleotide binding and hydrolysis play in the respective biological activities of p97-D1 and HslU.

The sensor-1 motif, which overlaps the "second region of homology" (9), contains a highly conserved "sensing" residue (Asn/Ser/Thr) found in close proximity to the gamma -phosphate (10). In Sec18p, mutation of Thr394 (the residue N-terminal to the sensing Asn) to proline results in a form of Sec18p that binds alpha -SNAP but is unable to stimulate ATP hydrolysis (27). The sensor-1 region extends through a short helix to a loop that is in close proximity to the nucleotide-binding site of an adjacent protomer in the hexamer. In most AAA proteins, this loop contains a highly conserved arginine residue (present in NSF-D1 but not NSF-D2). By analogy with G-proteins, this arginine residue has been proposed to be an "Arg finger" that can enhance the rate of hydrolysis of the nucleotide bound to an adjacent protomer (10, 28). In the p97-D1 structure, this arginine residue (Arg362) forms a salt bridge with Glu305 in the DEXX motif of the adjacent protomer. The side chain of p97-Arg359, also conserved among AAA proteins, is in close proximity to the beta -phosphate in the adjacent protomer. Either of these arginine residues could serve as a signal to the adjacent protomer upon nucleotide hydrolysis or release, providing a plausible mechanism for stimulation and cooperativity between hexamer subunits. Mutation of either of the equivalent residues in NSF-D1 (Arg385 or Arg388) prevents 20 S complex disassembly and does not affect either the basal or stimulated rates of hydrolysis.2 Mutation of the sensing Asn, the putative Arg finger, and other residues in close proximity results in the loss of protease activity in FtsH (28). As these mutants remain able to bind ATP, this phenotype has been attributed to the loss either of ATP hydrolysis or the coupling of hydrolysis to protease activity (28). In the HslUV chaperone-protease complex, the sensor-1 region of HslU is in direct contact with HslV (24) and is positioned to transmit changes in nucleotide state directly to its protease partner, HslV.

The sensor-2 motif, located at the N terminus of the third helix in the C-terminal subdomain (alpha 8 in NSF-D2) is in contact with the P-loop. This region has been proposed to be the primary site for transmitting conformational change from the nucleotide-binding site to the C-terminal subdomain (20, 22). The sensor-2 motif in HslU contacts both the P-loop and HslV in the HslUV complex (24) and is positioned to transmit changes upon nucleotide hydrolysis to both the C-terminal subdomain of HslU and HslV. In this light, it is interesting to note that in ClpA, ClpX, and Lon, which are also protease-associated AAA proteins, the C-terminal subdomains have been identified as the primary region for substrate recognition (reviewed in Ref. 29).

A number of structures now exist for HslU in the presence of different ATP analogues and in the absence of nucleotide (23, 24). When these structures are superimposed using their nucleotide-binding subdomains, a significant degree of flexibility is observed (23). Rigid body movements of the C-terminal subdomains (5-15°) occur about a pivot point in the intersubdomain linker. In the different crystal forms, the packing within the AAA hexamer changes significantly, and in some cases results in a break in the 6-fold symmetry of the hexamer. The changes in packing are probably due to relative movements of the N-terminal and C-terminal subdomains in protomers with different nucleotide occupancy (23). The observed differences may reflect crystal packing constraints rather than genuine conformational states but serve to illustrate that the hexameric interfaces may be sufficiently plastic to change during the catalytic cycle.

Recent electron microscopy data on p97 has presented additional possibilities for how changes in nucleotide state may be coupled to conformational change in AAA proteins (17, 26). In one model, derived from a three-dimensional EM reconstruction, the p97-D1 and p97-D2 hexamers are proposed to pack together in a tail-tail arrangement (17). Movements of the C-terminal subdomains of p97-D1 and p97-D2 would cause the p97-D1 and p97-D2 hexamers to rotate with respect to each other through a restricted angular range. These movements are proposed to be coupled to changes in nucleotide state in the p97-D1 and p97-D2 hexamers. The alternating movements of the two hexamers would cause p97 to act as a molecular ratchet and generate torsional force (17). It is unclear whether the tail-tail arrangement proposed for p97-D1 and p97-D2 is applicable to other AAA proteins containing tandem AAA domains. In NSF, for example, the sequence similarity between NSF-D1 and p97-D1 extends to the C-terminal residue observed in p97-D1 (Asn486 in NSF-D1). The N-terminal residue (Lys489) observed in the NSF-D2 crystal structure (21) is located on the "head" of the hexamer, is not influenced by lattice contacts, and leaves only two residues unaccounted for between the C terminus of NSF-D1 and the N terminus of NSF-D2. Furthermore, ATP hydrolysis by both AAA domains is not necessary for functional activity in NSF (7), PASI (30), or trypanosome p97 (31), ruling out the cooperation between D1 and D2 required for the proposed molecular ratchet mechanism. In the HslUV complex (26), the C terminus of HslU influences the active site architecture of HslV. If this mode of association between the protease and chaperone is conserved in ClpAP (where ClpA is an AAA protein containing tandem AAA domains and ClpP is the protease equivalent to HslV) then the two AAA domains of ClpA are unlikely to be arranged tail-tail.

In a separate study of p97 (26) little difference was seen between ADP- and ATP-bound forms. However, when nucleotide was removed from the samples, a large conformational change was observed, suggesting that nucleotide binding, rather than hydrolysis, could be the important step in the catalytic cycle (26). In support of this model, the largest intersubdomain movements in the different crystal forms of HslU are also seen between ATP-bound and nucleotide-free structures (23). It should be noted, however, that when the nucleotide-binding subdomains of NSF-D2 (ATP-bound) and p97-D1 (ADP-bound) are superimposed, the positions of the C-terminal helical subdomains are quite different (17). Given the differences in sequence between NSF-D2 and p97-D1, it is unclear whether this structural difference is a genuine reflection of changes upon nucleotide hydrolysis or a consequence of comparing domains from two different proteins.

    Mechanisms of SNARE Complex Disassembly
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However the SNARE complex is disassembled, force must be generated to separate the components. The physical mechanism by which NSF uses alpha -SNAP to disassemble SNARE complexes is not known, but the superhelical nature of the SNARE complex coiled-coil suggests that any mechanism of disassembly involves rotation of some kind. Physically this can only be achieved if part of at least one helix remains stationary while the remaining helices move, and two obvious possibilities can be considered. The first is that the membrane-anchored C terminus of the SNARE complex remains stationary while NSF and alpha -SNAP unravel the N-terminal ends. This mechanism is unlikely, as NSF can dissociate SNARE complexes formed only from the cytoplasmic portions of the SNARES, i.e. with no membrane anchors (32). Furthermore, the kinetics of 20 S complex disassembly are independent of whether the SNARE complex is in solution or located on membranes (33). The second possibility is that the N terminus of one or more of the components of the SNARE complex is attached directly to NSF, and movement of alpha -SNAPs bound to the remainder of the SNAREs relative to that anchor point unwinds the SNARE complex. This model requires the presence of a direct contact between components of the SNARE complex and NSF. NSF-alpha -SNAP can disassemble a core SNARE complex containing only the SNARE motifs of VAMP-2, syntaxin 1a, and SNAP-25 (32). The crystal structure of this core complex showed that the N termini of the syntaxin 1a and SNAP-25 N-terminal helices extend beyond the rest of the coiled-coil (34). These regions would then be the most likely points of contact with NSF. Although a direct interaction between SNAREs and NSF has never been detected, there is no reason to assume that it cannot take place in the context of the 20 S complex. This mechanism would operate equally well whether the SNAREs were attached to a membrane or in solution.

A further possibility for disassembly of the SNARE complex is that removal of part or all of one of the four helices in the coiled-coil would result in the spontaneous collapse of the remaining complex. In the case of the neuronal SNARE complex, the VAMP-2 and SNAP-25 SNARE motifs are unstructured in solution in the absence of syntaxin 1a (35). Given that NSF-alpha -SNAP is able to act on syntaxin 1a in the absence of other SNARE proteins (36), an NSF-induced conformational change imparted solely on syntaxin 1a may initiate melting of the VAMP and SNAP-25 helices, resulting in disassembly of the SNARE complex.

One of the crystal forms of NSF-N contained three protomers that were related by a pure 3-fold rotation axis (14). The dimensions of this trimeric NSF-N assembly were surprisingly concordant with those of the NSF-D2 hexamer and the "collar" region of the 20 S complex seen in electron micrographs (13). This, along with the proposed stoichiometry of 1 NSF hexamer:3 alpha -SNAP (33, 37) in the 20 S complex led to the suggestion that the trimer may represent a functional assembly. This presents the possibility that NSF works by using alternate sets of three NSF-N domains in a "3 in, 3 out" configuration. In this fashion, NSF would use the sleeve of alpha -SNAP molecules to apply torsion to the SNARE complex and unwind it (14). Alternatively, NSF may use the alternate sets of NSF-N domains to pull one end of alpha -SNAP toward the outside of the NSF hexamer and unravel the SNARE coiled-coil.

The observation of the central pore in NSF has led to speculation about its possible functional role (21, 34), particularly when considered in the context of other AAA proteins. The protease-associated AAA chaperones feed unfolded proteins through the central pore of the AAA hexamer to meet their fate in the active site of the protease (29). This type of activity is analogous to that seen in some hexameric DNA helicases which unravel DNA by passing one strand of the DNA duplex through their central pore while excluding the complimentary strand (38). Given the dimensions of the central pore in the NSF-D2 hexamer it is possible that an unfolded peptide chain could pass through the pore and be separated from the remainder of the SNARE complex. This model presents several problems. It is unclear which of the SNARE proteins would pass through the central pore. The syntaxin family of t-SNAREs has a conserved globular N-terminal domain with dimensions significantly larger than the NSF-D2 pore. The same problem applies to some v-SNAREs (e.g. rSec22, VAMP-7) that contain a N-terminal globular domain (39). SNAP-25, which contributes two helices to the four-helix bundle SNARE complex, would also have difficulties passing through the NSF-D2 pore because of the membrane association of the linker region between its two SNARE motifs. Sec9p, the yeast orthologue of SNAP-25, has a large domain N-terminal to the two SNARE motifs. If the NSF-D1 ring operates by a pore insertion mechanism, large scale remodeling of the hexamer would have to take place, which is not consistent with electron microscopic images.

    Concluding Remarks
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NSF was first identified 12 years ago (40). Since then a large amount of information has been accumulated about its biological activity in many different species. Despite this, remarkably little is known about how NSF transforms the energy of ATP hydrolysis into the mechanical work needed to disassemble SNARE complexes. Many important lessons can be learned through comparison with other proteins that contain AAA domains, but specific questions about NSF remain unanswered. How is ATP hydrolysis actually coupled to SNARE complex disassembly, and how many ATPs are required? How do the components of the 20 S complex interact? How do NSF and alpha -SNAP act upon complexes of SNARE proteins from different transport steps? Ongoing structural, biochemical, and biophysical studies of NSF and related proteins should provide answers to these questions over the next few years.

    ACKNOWLEDGEMENTS

We thank Xiaodong Zhang and Paul Freemont for providing the coordinates of p97-ND1.

    FOOTNOTES

* This minireview will be reprinted in the 2001 Minireview Compendium, which will be available in December, 2001.

§ Funded by a Wellcome Trust International Prize Traveling Fellowship.

|| Supported by NHLBI, National Institutes of Health.

** Supported in part by Grant MH58570 from the National Institute of Mental Health. To whom correspondence should be addressed. Tel.: 650-725-4623; Fax: 650-723-8464; E-mail: bill.weis@stanford.edu.

Published, JBC Papers in Press, April 11, 2001, DOI 10.1074/jbc.R100013200

2 E. A. Matveeva and S. W. Whiteheart, unpublished data.

3 See supplementary information associated with Ref. 17.

    ABBREVIATIONS

The abbreviations used are: SNARE, soluble NSF attachment protein receptors; v-SNARE, vesicle SNARE; t-SNARE, target SNARE; SNAP, soluble NSF attachment protein; NSF, N-ethylmaleimide-sensitive factor; EM, electron micrograph; AMP-PNP, adenosine 5'-(beta ,gamma -imino)triphosphate; ATPgamma S, adenosine 5'-3-O-(thio)triphosphate; AAA, ATPases associated with various cellular activities; VAMP, vesicle-associated membrane protein.

    REFERENCES
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1. Jahn, R., and Südhof, T. C. (1999) Annu. Rev. Biochem. 68, 863-911[CrossRef][Medline] [Order article via Infotrieve]
2. Wilson, D. W., Whiteheart, S. W., Wiedmann, M., Brunner, M., and Rothman, J. E. (1992) J. Cell Biol. 117, 531-538[Abstract]
3. Tagaya, M., Wilson, D. W., Brunner, M., Arango, N., and Rothman, J. E. (1993) J. Biol. Chem. 268, 2662-2666[Abstract/Free Full Text]
4. Morgan, A., Dimaline, R., and Burgoyne, R. D. (1994) J. Biol. Chem. 269, 29347-29350[Abstract/Free Full Text]
5. Matveeva, E., and Whiteheart, S. W. (1998) FEBS Lett. 435, 211-214[CrossRef][Medline] [Order article via Infotrieve]
6. Nagiec, E. E., Bernstein, A., and Whiteheart, S. W. (1995) J. Biol. Chem. 270, 29182-29188[Abstract/Free Full Text]
7. Whiteheart, S. W., Rossnagel, K., Buhrow, S. A., Brunner, M., Jaenicke, R., and Rothman, J. E. (1994) J. Cell Biol. 126, 945-954[Abstract]
8. Matveeva, E. A., He, P., and Whiteheart, S. W. (1997) J. Biol. Chem. 272, 26413-26418[Abstract/Free Full Text]
9. Patel, S., and Latterich, M. (1998) Trends Cell Biol. 8, 65-71[CrossRef][Medline] [Order article via Infotrieve]
10. Neuwald, A. F., Aravind, L., Spouge, J. L., and Koonin, E. V. (1999) Genome Res. 9, 27-43[Abstract/Free Full Text]
11. Vale, R. D. (2000) J. Cell Biol. 150, F13-F20[Abstract/Free Full Text]
12. Hanson, P. I., Roth, R., Morisaki, H., Jahn, R., and Heuser, J. E. (1997) Cell 90, 523-535[Medline] [Order article via Infotrieve]
13. Hohl, T. M., Parlati, F., Wimmer, C., Rothman, J. E., Sollner, T. H., and Engelhardt, H. (1998) Mol. Cell 2, 539-548[Medline] [Order article via Infotrieve]
14. May, A. P., Misura, K. M. S., Whiteheart, S. W., and Weis, W. I. (1999) Nat. Cell Biol. 1, 175-182[CrossRef][Medline] [Order article via Infotrieve]
15. Yu, R. C., Jahn, R., and Brunger, A. T. (1999) Mol. Cell 4, 97-107[Medline] [Order article via Infotrieve]
16. Babor, S. M., and Fass, D. (1999) Proc. Natl. Acad. Sci. U. S. A. 96, 14759-14764[Abstract/Free Full Text]
17. Zhang, X., Shaw, A., Bates, P. A., Newman, R. H., Gowen, B., Orlova, E., Gorman, M. A., Kondo, H., Dokurno, P., Lally, J., Leonard, G., Meyer, H., van Heel, M., and Freemont, P. S. (2000) Mol. Cell 6, 1473-1484[Medline] [Order article via Infotrieve]
18. Coles, M., Diercks, T., Liermann, J., Groger, A., Rockel, B., Baumeister, W., Koretke, K. K., Lupas, A., Peters, J., and Kessler, H. (1999) Curr. Biol. 9, 1158-1168[CrossRef][Medline] [Order article via Infotrieve]
19. Barnard, R. J. O., Morgan, A., and Burgoyne, R. D. (1996) Mol. Biol. Cell 7, 693-701[Abstract]
20. Lenzen, C. U., Steinmann, D., Whiteheart, S. W., and Weis, W. I. (1998) Cell 94, 525-536[Medline] [Order article via Infotrieve]
21. Yu, R. C., Hanson, P. I., Jahn, R., and Brünger, A. T. (1998) Nat. Struct. Biol. 5, 803-811[CrossRef][Medline] [Order article via Infotrieve]
22. Guenther, B., Onrust, R., Sali, A., O'Donnell, M., and Kuriyan, J. (1997) Cell 91, 335-345[Medline] [Order article via Infotrieve]
23. Bochtler, M., Hartmann, C., Song, H. K., Bourenkov, G. P., Bartunik, H. D., and Huber, R. (2000) Nature 403, 800-805[CrossRef][Medline] [Order article via Infotrieve]
24. Sousa, M. C., Trame, C. B., Tsuruta, H., Wilbanks, S. M., Reddy, V. S., and McKay, D. B. (2000) Cell 103, 633-643[Medline] [Order article via Infotrieve]
25. Rockel, B., Walz, J., Hegerl, R., Peters, J., Typke, D., and Baumeister, W. (1999) FEBS Lett. 451, 27-32[CrossRef][Medline] [Order article via Infotrieve]
26. Rouiller, I., Butel, V. M., Latterich, M., Milligan, R. A., and Wilson-Kubalek, E. M. (2000) Mol. Cell 6, 1485-1490[Medline] [Order article via Infotrieve]
27. Steel, G. J., Harley, C., Boyd, A., and Morgan, A. (2000) Mol. Biol. Cell 11, 1345-1356[Abstract/Free Full Text]
28. Karata, K., Inagawa, T., Wilkinson, A. J., Tatsuta, T., and Ogura, T. (1999) J. Biol. Chem. 274, 26225-26332[Abstract/Free Full Text]
29. Schmidt, M., Lupas, A. N., and Finley, D. (1999) Curr. Opin. Chem. Biol. 3, 584-591[CrossRef][Medline] [Order article via Infotrieve]
30. Hohfeld, J., Mertens, D., Wiebel, F. F., and Kunau, W. H. (1992) in Membrane Biogenesis and Protein Targeting (Neupert, W. , and Lill, R., eds) , pp. 185-207, Elsevier Science Publishing Co., Inc., New York
31. Lamb, J. R., Fu, V., Wirtz, E., and Bangs, J. D. (2001) J. Biol. Chem. 276, in press
32. Fasshauer, D., Otto, H., Eliason, W. K., Jahn, R., and Brunger, A. T. (1997) J. Biol. Chem. 272, 28036-28041[Abstract/Free Full Text]
33. Swanton, E., Bishop, N., Sheehan, J., High, S., and Woodman, P. (2000) J. Cell Sci. 113, 1783-1791[Abstract/Free Full Text]
34. Sutton, R. B., Fasshauer, D., Jahn, R., and Brünger, A. T. (1998) Nature 395, 347-353[CrossRef][Medline] [Order article via Infotrieve]
35. Fiebig, K. M., Rice, L. M., Pollock, E., and Brunger, A. T. (1999) Nat. Struct. Biol. 6, 117-123[CrossRef][Medline] [Order article via Infotrieve]
36. Hanson, P. I., Otto, H., Barton, N., and Jahn, R. (1995) J. Biol. Chem. 270, 16955-16961[Abstract/Free Full Text]
37. Hayashi, T., Yamasaki, S., Nauenburg, S., Binz, T., and Niemann, H. (1995) EMBO J. 14, 2317-2325[Abstract]
38. Egelman, E. H. (1998) J. Struct. Biol. 24, 123-128
39. Gonzalez, L. C., Jr., Weis, W. I., and Scheller, R. H. (2001) J. Biol. Chem. 276, in press
40. Block, M. R., Glick, B. S., Wilcox, C. A., Wieland, F. T., and Rothman, J. E. (1988) Proc. Natl. Acad. Sci. U. S. A. 85, 7852-7856[Abstract]
41. Rice, L. M., and Brunger, A. T. (1999) Mol. Cell 4, 85-95[Medline] [Order article via Infotrieve]
42. Esnouf, R. M. (1997) J. Mol. Graph. 15, 133-138
43. Merritt, E. A., and Bacon, D. J. (1997) Methods Enzymol. 277, 505-524


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