NIH News Release
National Institute of Mental Health

Thursday, April 6, 2000
Jules Asher
(301) 443-4536

Molecular Minuet of Membrane Fusion Proteins Unveiled

The vision of chemical messengers crossing the synapse and binding to receptors may command the spotlight. But to solve the mystery of how brain cells communicate, neuroscientists are increasingly peeking behind the scenes — at what's going on inside the secreting cell. There, electrical impulses propel vesicles, tiny spheres containing neurotransmitters, into the cell wall. Just before a neuron sprays a neurotransmitter into the synapse, mercurial molecules perform a secret dance that fuses the vesicle membrane with the cell wall membrane. Proteins change their shapes and locales, as their Velcro-like extensions intertwine to conjure a momentary dragon. This highly charged core complex melds the membranes, then vanishes. In the March 23, 2000 issue of Nature, NIMH grantees Drs. Richard Scheller, William Weis and colleagues at Stanford University reveal, for the first time, the atom-by-atom, 3-D structure and cagey choreography of a lead hoofer in this act, the Sec1-syntaxin1a complex.

Likened to soap bubbles merging, or bubbles bursting at the surface of boiling water, membrane fusion (*RealPlayer format) has attracted heightened interest among neuroscientists in recent years. The family of proteins involved, SNARES (Soluble NSF Attachment protein REceptor), have been conserved through evolution, performing similar functions even in primitive organisms like yeast and fruit flies. Some of the fusion proteins are embedded in the vesicle and the cell wall membranes; others float freely within the nerve terminal, or axon.

The entire neurotransmission performance — from electrical signal to receptor binding — lasts less than a thousandth of a second, and is repeated billions of times daily in each of the human brain's 100 billion neurons. So membrane fusion mechanisms may hold clues about what goes wrong in disorders of thinking, learning and memory, including schizophrenia and other mental illnesses thought to involve disturbances in neuronal communication.

Scheller, Weis and doctoral student Kira Misura were especially interested in a complex created when two proteins bind together: the free-floating SNARE regulator nsec1, and the target membrane's sharply-bent, helical SNARE, syntaxin1. Formation of the complex is the first act in the membrane fusion ballet. To reveal the complex's secret moves, the researchers examined its crystalline structure using a type of subatomic particle accelerator called a synchrotron. The mile-long accelerator generates very intense x-ray beams used to resolve objects as small as atoms, and capture snapshots of the shapes of molecules at different stages of chemical reactions.

Based on the revealed physical properties, as well as genetic and biochemical evidence, the investigators propose that sec1/syntaxin1 complex communicates with Rab, a G protein on the vesicle membrane, and Rab-E, a free-floating cousin, to recognize and guide the neurotransmitter-filled sac to its appointed fusion site on the target membrane. They further propose that syntaxin1a and Sec1 collaborate with VAMP, a velcro-like vesicle membrane protein, and SNAP-25, a counterpart on the cell's membrane, to bring about fusion itself.

In this model, nSec1 performs a "chaperone-like" role for syntaxin1a, first shielding it from other proteins, and later facilitating its interactions with them. When in its protective posture, nSec1 renders its partner physically incapable of forming stable relationships with other proteins.

The steps to the dance are as follows (See diagram of model below):

A nSec1 binds tightly to, and cloaks, syntaxin1a, forming a complex that serves as a recognition site for an arriving vesicle.

B The vesicle's Rab and/or Rab-E proteins recognize the complex, signaling nSec1 to change its shape and move outward, uncovering a key binding region of syntaxin1a, and destabilizing it. Residues from the suddenly exposed binding region signal SNAP-25 to begin making the core fusion complex, using nSec1 as a platform.

C Syntaxin1a's sharply bent end section moves away from its key binding region, making way for SNAP-25 and VAMP to move in and bind.

D VAMP/SNAP-25/syntaxin1a combine to form a long, straight, helical bundle, the core complex. This pulls the vesicle and target membranes together, fusing them, and releasing the neurotransmitter. After fusion, an enzyme breaks down the complex and the SNAREs are recycled for a repeat performance.

The study was conducted on mammalian proteins, but also took advantage of forms of nSec1 and syntaxin1a found in mutant fruit flies. It is the first study by a NIMH-funded team to resolve the changing atomic structure of a protein.

The National Institute of Mental Health (NIMH) is part of the National Institutes of Health (NIH), the Federal Government's primary agency for biomedical and behavioral research. NIH is a component of the U.S. Department of Health and Human Services.

'steps to the dance' of membrane fusion

Kira M.S. Misura, Richard H. Scheller, William I. Weis, "Three-dimensional structure of the neuronal-Sec1syntaxin 1a complex," Nature, March 23, 2000. 404, 355-362.