Andreas Bracher

The Matrix Protein Vp40 from Ebola Virus Octamerizes Into Pore-Like Structures with Specific RNA Binding Properties

Abstract The Ebola virus membrane-associated matrix protein VP40 is thought to be crucial for assembly and budding of virus particles. Here we present the crystal structure of a disk-shaped octameric form of VP40 formed by four antiparallel homodimers of the N-terminal domain. The octamer binds an RNA triribonucleotide containing the sequence 5′-U-G-A-3′ through its inner pore surface, and its oligomerization and RNA binding properties are facilitated by two conformational changes when compared to monomeric VP40. The selective RNA interaction stabilizes the ring structure and confers in vitro SDS resistance to octameric VP40. SDS-resistant octameric VP40 is also found in Ebola virus-infected cells, which suggests that VP40 has an additional function in the life cycle of the virus besides promoting virus assembly and budding off the plasma membrane.

Crystal structure of the GABAA‐receptor‐associated protein, GABARAP

The GABAA‐receptor‐associated protein (GABARAP) is a member of a growing family of intracellular membrane trafficking and/or fusion proteins and has been implicated in plasma membrane targeting and/or recycling of GABAA receptors. GABARAP is localized on intracellular membranes such as the trans‐Golgi network, binds to the γ 2 subunit of GABAA receptors and interacts with microtubules and the N‐ethylmaleimide‐sensitive factor. We report the X‐ray crystal structure of mammalian GABARAP at 2.0 Å resolution. GABARAP consists of an N‐terminal basic helical region, which has been implicated in tubulin binding, and a core structure with a conserved ubiquitin‐like fold. Consistent with the high extent of sequence conservation among GABARAP homologues from plants to mammals, one face of the core structure is absolutely conserved while the opposite face shows considerable divergence. These features are in agreement with the conserved surface mediating protein–protein interactions shared by all members of the family, whereas the non‐conserved surface region may play specific roles, such as docking to particular membrane receptors.

The X-ray crystal structure of neuronal Sec1 from squid sheds new light on the role of this protein in exocytosis.

BACKGROUND Sec1-like molecules have been implicated in a variety of eukaryotic vesicle transport processes including neurotransmitter release by exocytosis. They regulate vesicle transport by binding to a t-SNARE from the syntaxin family. This process is thought to prevent SNARE complex formation, a protein complex required for membrane fusion. Whereas Sec1 molecules are essential for neurotransmitter release and other secretory events, their interaction with syntaxin molecules seems to represent a negative regulatory step in secretion. RESULTS Here we report the X-ray crystal structure of a neuronal Sec1 homologue from squid, s-Sec1, at 2.4 A resolution. Neuronal s-Sec1 is a modular protein that folds into a V-shaped three-domain assembly. Peptide and mutagenesis studies are discussed with respect to the mechanism of Sec1 regulation. Comparison of the structure of squid s-Sec1 with the previously determined structure of rat neuronal Sec1 (n-Sec1) bound to syntaxin-1a indicates conformational rearrangements in domain III induced by syntaxin binding. CONCLUSIONS The crystal structure of s-Sec1 provides the molecular scaffold for a number of molecular interactions that have been reported to affect Sec1 function. The structural differences observed between s-Sec1 and the structure of a rat n-Sec1-syntaxin-1a complex suggest that local conformational changes are sufficient to release syntaxin-1a from neuronal Sec1, an active process that is thought to involve additional effector molecule(s).

Hsp90 structure: when two ends meet

The crystal structure of the full-length yeast Hsp90 sheds new light on the conformational cycle and the functional regulation of this important molecular chaperone.

Crystallization and preliminary X-ray analysis of squid neuronal Sec1.

Sec1 protein family members are involved in the regulation of all intracellular SNARE-mediated (SNARE = soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor) vesicle-fusion processes in a step preceding membrane fusion and have been shown to interact with t-SNAREs. To better understand the structural basis and the role of Sec1 in the regulation of the SNARE-complex formation, neuronal Sec1 from the squid Loligo pealei has been expressed and crystallized; this invertebrate protein shows a high sequence homology to the human neuronal Sec1, Munc18a. Here, the production of diffraction-quality native crystals, which belong to space group P3(1)21 and diffract to 3.3 A resolution, is described. In addition, selenomethionyl n-Sec1 crystals in space groups P3(1)21 and P2(1) have been generated. Preliminary analysis of the monoclinic space group indicates that these crystals diffract to a resolution higher than 2.5 A.

Studies on the Reaction Mechanism of GTP Cyclohydrolase I

GTP cyclohydrolase I catalyzes a ring expansion affording dihydroneopterin triphosphate from GTP (1, 2). The reaction involves the release of C-8 of GTP as formate. The catalytic domain of the human enzyme shows 37% identity with that of Escherichia coli. Both enzymes are toroid-shaped homodecamers with d5 symmetry and were recently shown to contain a catalytically essential zinc ion at each of the 10 equivalent active sites (Figure 1) (3, 4).