determined motional amplitudes1 are in gen-eral agreement with the known amplitudes of picosecond dynamics previously obtained by NMR7. The newly determined amplitudes also agree well with a molecular-dynamics simula-tion of the protein performed by the authors to validate their results.

More excitingly, using their technique, Salmon et al. observed motions that occur on a longer timescale (> 400 nanoseconds) than could be accessed by the molecular-dynamics simulation, specifically for a β-turn region and for a loop that lies close to ubiquitin’s protein-binding interface. These motions could not have been observed using previously available NMR methods. The authors’ results reinforce a picture of ubiquitin as a protein that has a rigidly packed, stable fold in which most of the flexibility is concentrated in regions responsible for its functional roles — protein recognition and assembly into covalently linked polyubiq-uitin chains8–10 that serve as cellular signals. This defuses a long-standing controversy about whether globular, non-enzymatic pro-teins such as ubiquitin undergo any motions on timescales longer than nanoseconds.

Salmon and colleagues’ method1 opens the door to studies of other proteins that are expected to be more flexible than ubiquitin. At present, the main obstacle to using the authors’ technique is that many different solutions of the protein must be prepared, each providing envi-ronments that cause the protein molecules to align differently. This can be quite challenging. Nevertheless, the method enables a straight-forward average of the atomic coordinates of proteins to be recorded for movements occur-ring over the full submillisecond time scale, which clearly has important implications. Knowledge of the extent of protein movements could enable the determination of ensembles of structures that represent the conformational fluctuations of proteins, help to guide the design of ligands, and advance our understanding of protein catalysis and allostery (the regulation of protein activity by ligand binding to sites other than the functional binding site).

Dynamics is often used to explain apparent discrepancies between structural models of proteins and their known functions, yet there are few cases in which direct links between dynamics and function have been made. As experimental techniques for determining dynamics continue to improve, we can look forward to gaining deeper insight into the question of whether dynamics is more often essential or extraneous to protein function. ■

Joel R. Tolman is in the Department of Chemistry,

Johns Hopkins University, 3400 North Charles

Street, Baltimore, Maryland 21218, USA.

e-mail: tolman@jhu.edu

1. Salmon, L. et al. Angew. Chem. Int. Edn 48, 4154–4157

(2009).

2. Perutz, M. F. & Mathews, F. S. J. Mol. Biol. 21, 199–202

(1966).

3. Bustamante, C. Annu. Rev. Biochem. 77, 45–50 (2008).

4. Mittermaier, A. & Kay, L. E. Science 312, 224–228 (2006).

5. Meiler, J., Prompers, J. J., Peti, W., Griesinger, C.

& Brüschweiler, R. J. Am. Chem. Soc. 123, 6098–6107

(2001).

6. Tolman, J. R. J. Am. Chem. Soc. 124, 12020–12030 (2002).

7. Tjandra, N., Feller, S. E., Pastor, R. W. & Bax, A. J. Am. Chem.

Soc. 117, 12562–12566 (1995).

8. Briggman, K. B. & Tolman, J. R. J. Am. Chem. Soc. 125, 10164–10165 (2003).

9. Lange, O. F. et al. Science 320, 1471–1475 (2008).

10. Peti, W., Meiler, J., Brüschweiler, R. & Griesinger, C.

J. Am. Chem. Soc. 124, 5822–5833 (2002).

GRANULAR MEDIA

Structures in sand streamsDetlef Lohse and Devaraj van der Meer

An ingenious experiment that involves dropping a costly, high-speed video camera from a height of several metres reveals how free-falling streams of granular matter, such as sand, break up into grain clusters.

It is common knowledge that when a stone is thrown into a pond, a jet of water shoots upwards. High-speed video imaging and even snapshots taken with short exposure times reveal that the jet breaks up into drop-let patterns. Such patterns have inspired artists such as Andrew Davidhazy. What is both remarkable and intriguing is that very similar arrangements are observed in the streams that emerge when a stone is dropped into loose, fine sand1. The common view, which we have also adhered to, about the origin of this striking, liquid-like behaviour of streams of sand or other granular materials has been that the inelastic nature of collisions between the grains causes them to cluster together into larger structures2–4. But on page 1110 of this issue, Royer and colleagues5 show that this view is wrong, and that forces hitherto believed to be too small to cause the clustering are at work.

The formation of droplets in liquid jets (Fig. 1a) is caused by the Rayleigh–Plateau instability, which is driven by surface tension (the force that causes a liquid droplet to keep its shape). The understanding of this instability was pivotal to the conceptual development of hydrodynamics in the late nineteenth century by Joseph Plateau, Lord Rayleigh and their successors, and has since become textbook knowledge6. In contrast to liquid jets, granu-lar streams of matter (Fig. 1b) were believed to lack surface tension. After all, granular matter such as sand is defined as a collection of grains that exert no forces on each other, with the exception of repulsive forces on collision7. It is thus surprising that streams of granular matter break up into structures that are similar to those of liquid streams.

Now Royer and colleagues5 demonstrate that, in granular matter, tiny nanometre-range forces between the grains lead to a minute effec-tive surface tension that, despite being 100,000 times smaller than that in water, can explain the clustering of grains in the jet. The researchers achieved this using an ingenious combination of nanometre-scale atomic-force-microscopy measurements of the forces between the sand grains with metre-scale tracking of the

evolution of the granular streams. The track-ing involved letting sand (and other granular material) stream out of a funnel and following its clustering dynamics, for almost a second, in the stream’s co-moving frame of reference — that is, using a high-speed video camera that moved with the stream. This means that the researchers had to drop their US$80,000 high-speed camera from a height of several metres, a heart-stopping prospect — even though the impact of the camera on the ground was adequately buffered.

In their experiment, Royer et al.5 observe a direct correlation between nanometre-range cohesive forces between the grains and the evolution of the jet structures within the first few metres of free fall. They find that modify-ing the nanometre-range forces — by varying the strength of intergrain cohesion through changes in the grains’ surface roughness, the humidity or by using different materi-als — directly affects the break-up dynamics of the streams into clusters. In particular, the authors note that suppression of the nano-metre-scale cohesion causes the clustering to vanish.

We would like to illustrate this striking phenomenon with two analogies. The first relates to the shapes of the grain clusters. As the authors5 themselves note, the shapes of the clusters, including the double-cone necks at rupture, resemble the droplets that form in liquid nanojets and which were first found in molecular-dynamics studies that simulated injection of molecular fluids into a vacuum through a nanoscale nozzle8. Such studies show that the small number of molecules and thermal fluctuations in the nanojets lead to irregulari-ties in the cluster shapes, very similar to those found in Royer and colleagues’ granular jets, in which the number of sand grains is also small when compared with the huge number of molecules in macroscopic liquid jets.

The second parallel that can be made is with the emergence of planetary systems from dust grains in circumstellar gas disks. In this case, the first step is the formation of metre-sized boulders, but how this process continues to form kilometre-sized planetesimals is an

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unsolved problem9, given that the (gravita-tional) attractive force between the boulders is so weak. However, Royer and colleagues’ work teaches us that even minute forces can drive the formation of structures if the system is given enough time.

Traditionally, and in contrast to powders, one of the defining properties of granu-lar matter has been the absence of cohesive forces between the grains7. But Royer and colleagues’ observation of clustering in granu-lar jets implies, conversely to what scientists have previously believed, that the distinction between powders and granular matter is much less clear-cut. Indeed, when velocity differences between the grains are small and timescales are considerable, even large grains can behave as powders.

The consequences of small, attractive forces may reach beyond the free-falling granu-lar jets1. For example, the structures formed in the sand ‘splashes’ that are blown away after the impact of heavy objects on soft, fine sand1 could presumably be correlated

Figure 1 | Rupture of streams. a, In a process caused by surface tension, a water stream breaks up into droplets (image taken from ref. 10). b, Similarly, a stream of granular media such as a sand jet — which forms, for example, after the impact of an object on loose, fine sand — breaks up into clusters of grains (image taken from ref. 1). Royer and colleagues5 show that, in granular media, the rupture and clustering process is driven by nanometre-range forces between the grains that lead to an effective surface tension that is 100,000 times weaker than in ordinary liquids.

a b with the same type of cohesive force. Future studies are needed to investigate whether such a correlation exists, just as Royer and colleagues have done in this pioneering study of granular jets. ■

Detlef Lohse and Devaraj van der Meer are in

the Physics of Fluids Group, Department of

Science and Technology, J. M. Burgers Center,

and at the Impact and Mesa+ Institutes,

University of Twente, PO Box 217, 7500 AE

Enschede, the Netherlands.

e-mail: d.lohse@utwente.nl

CELL BIOLOGY

A score for membrane fusionRuth N. Collins and Joshua Zimmerberg

Intracellular membrane fusion has been mimicked in vitro using a mix of 17 purified proteins and lipid bilayers. This technical tour de force allows the study of how cells orchestrate and perform such fusion events.

Molecules in living organisms are constantly being replaced, yet cellular structures can maintain their identity for a lifetime1. About half of all biological processes involve mem-brane proteins, which must be delivered, and eventually removed, with great accuracy to regulate the constancy of structural identity. This delivery and removal is mediated by the membrane-bound organelles of eukaryotic cells, which communicate with each other by budding off vesicles and other transport packages, which travel to, and fuse with, tar-get membranes. The organization and regula-tion of membrane-fusion reactions, then, are crucial for virtually every membrane-bound biological process.

But how is fusion of internal membranes achieved? To understand these mechanisms, researchers have strived to devise in vitro systems that mimic fusion events in vivo. In this issue (page 1091), Ohya et al.2 describe an enormous technical accomplishment — the self-assembly of 17 individual purified pro-teins into a fusion ‘machine’ whose activity and regulation recapitulate those of membrane fusion in an intact cell.

Two decades of investigation into mem-brane fusion have given us a set of key players. These include the SNARE family of membrane proteins and the Rab proteins, a subfamily of the Ras superfamily of GTPase enzymes. Mutations or pharmacological treatment of any of these components block fusion in vivo3. The membrane-bound SNARE proteins link the two membranes destined for fusion. They do this by interacting with partner SNARE proteins on the opposing membrane to form a stable, four-coiled bundle consist-ing of helices from several individual SNARE proteins4 (Fig. 1, overleaf).

Rab GTPases, such as Rab5, are found both

on membranes and in the cytosol. Rab pro-teins alternate between GDP-bound and GTP-bound forms, a switch controlled by proteins called guanine-nucleotide exchange factors. Like that of other members of the Ras super-family, the nucleotide-dependent switch of Rab proteins is thought to control a downstream catalytic process. Rab proteins are involved in such pro cesses as the regulation of organelle transport, the tethering of membranes before fusion, and the control of SNARE function5.

Each of these protein families is served by a constellation of accessory factors; these include NSF, an ATPase enzyme that helps to recycle the SNARE protein complex; the Sec1/Munc18 (SM) proteins, which together with SNAREs are essential for fusion; and various effec-tor proteins that interact with GTP-bound, active Rab4,5.

SNARE complexes are structurally similar to viral envelope proteins known to catalyse membrane fusion6, so a paradigm emerged casting these rod-shaped helical bundles into the central role of ‘minimal fusion machine’, with all other proteins assigned to supporting roles such as regulation of the SNARE com-plex7. However, direct tests comparing the contributions of SNAREs alone with SNAREs plus all the other proteins essential for fusion have not been performed because of the com-plexities of assembling microgram quantities of membrane proteins in defined lipid environ-ments together with other fragile purified proteins — requirements that stretch the limits of current biochemical technologies.

Ohya et al.2 overcome these obstacles, and reconstitute membrane fusion in vitro using physiological concentrations of 17 proteins derived from human membrane-bound organelles called endosomes, together with endosomal lipids. They quantify fusion by

a, C

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2. Jaeger, H. M., Nagel, S. R. & Behringer, R. P. Rev. Mod. Phys.

68, 1259–1273 (1996).

3. Kadanoff, L. Rev. Mod. Phys. 71, 435–444 (1999).

4. Goldhirsch, I. Annu. Rev. Fluid Mech. 35, 267–293 (2003).

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6. de Gennes, P.-G., Brochard-Wyart, F. & Quere, D. Capillarity

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