Crystalline ribonuclease A loses function below the dynamical transition at 220 K.

When the dynamic properties of many different proteins are plotted as a function of temperature, biphasic behaviour is observed, with a broad transition centred around 220 K. Atomic mean-square displacements from X-ray crystallography and M?ssbauer scattering show this behaviour, as do electron transfer rates and dynamic information from inelastic neutron scattering. Molecular dynamics simulations over a range of temperatures also exhibit a transition at about 220 K: high-temperature atomic fluctuations are dominated by anharmonic collective motions of bonded and nonbonded groups of atoms, but below 220 K the predominant dynamic behaviour is harmonic vibration of individual atoms. Here we show by high-resolution X-ray diffraction that crystalline ribonuclease A does not bind substrate or inhibitor at 212 K but will bind either rapidly at 228 K. Once bound at the higher temperature, inhibitor cannot be washed off after the enzyme is cooled to below the transition temperature. These results suggest that enzyme flexibility is required for catalytic function.     

Release of methane from a volcanic basin as a mechanism for initial Eocene global warming

A 200,000-yr interval of extreme global warming marked the start of the Eocene epoch about 55 million years ago. Negative carbon- and oxygen-isotope excursions in marine and terrestrial sediments show that this event was linked to a massive and rapid (approximately 10,000 yr) input of isotopically depleted carbon. It has been suggested previously that extensive melting of gas hydrates buried in marine sediments may represent the carbon source and has caused the global climate change. Large-scale hydrate melting, however, requires a hitherto unknown triggering mechanism. Here we present evidence for the presence of thousands of hydrothermal vent complexes identified on seismic reflection profiles from the V?ring and M?re basins in the Norwegian Sea. We propose that intrusion of voluminous mantle-derived melts in carbon-rich sedimentary strata in the northeast Atlantic may have caused an explosive release of methane--transported to the ocean or atmosphere through the vent complexes--close to the Palaeocene/Eocene boundary. Similar volcanic and metamorphic processes may explain climate events associated with other large igneous provinces such as the Siberian Traps (approximately 250 million years ago) and the Karoo Igneous Province (approximately 183 million years ago).     

Mutations in EFHC1 cause juvenile myoclonic epilepsy

Juvenile myoclonic epilepsy (JME) is the most frequent cause of hereditary grand mal seizures. We previously mapped and narrowed a region associated with JME on chromosome 6p12-p11 (EJM1). Here, we describe a new gene in this region, EFHC1, which encodes a protein with an EF-hand motif. Mutation analyses identified five missense mutations in EFHC1 that cosegregated with epilepsy or EEG polyspike wave in affected members of six unrelated families with JME and did not occur in 382 control individuals. Overexpression of EFHC1 in mouse hippocampal primary culture neurons induced apoptosis that was significantly lowered by the mutations. Apoptosis was specifically suppressed by SNX-482, an antagonist of R-type voltage-dependent Ca(2+) channel (Ca(v)2.3). EFHC1 and Ca(v)2.3 immunomaterials overlapped in mouse brain, and EFHC1 coimmunoprecipitated with the Ca(v)2.3 C terminus. In patch-clamp analysis, EFHC1 specifically increased R-type Ca(2+) currents that were reversed by the mutations associated with JME.     

Large expansion of the ATTCT pentanucleotide repeat in spinocerebellar ataxia type 10

Spinocerebellar ataxia type 10 (SCA10; MIM 603516; refs 1,2) is an autosomal dominant disorder characterized by cerebellar ataxia and seizures. The gene SCA10 maps to a 3.8-cM interval on human chromosome 22q13-qter (refs 1,2). Because several other SCA subtypes show trinucleotide repeat expansions, we examined microsatellites in this region. We found an expansion of a pentanucleotide (ATTCT) repeat in intron 9 of SCA10 in all patients in five Mexican SCA10 families. There was an inverse correlation between the expansion size, up to 22.5 kb larger than the normal allele, and the age of onset (r2=0.34, P=0.018). Analysis of 562 chromosomes from unaffected individuals of various ethnic origins (including 242 chromosomes from Mexican persons) showed a range of 10 to 22 ATTCT repeats with no evidence of expansions. Our data indicate that the new SCA10 intronic ATTCT pentanucleotide repeat in SCA10 patients is unstable and represents the largest microsatellite expansion found so far in the human genome.     

Crystal structure of the human beta2 adrenergic G-protein-coupled receptor

Structural analysis of G-protein-coupled receptors (GPCRs) for hormones and neurotransmitters has been hindered by their low natural abundance, inherent structural flexibility, and instability in detergent solutions. Here we report a structure of the human beta2 adrenoceptor (beta2AR), which was crystallized in a lipid environment when bound to an inverse agonist and in complex with a Fab that binds to the third intracellular loop. Diffraction data were obtained by high-brilliance microcrystallography and the structure determined at 3.4 A/3.7 A resolution. The cytoplasmic ends of the beta2AR transmembrane segments and the connecting loops are well resolved, whereas the extracellular regions of the beta2AR are not seen. The beta2AR structure differs from rhodopsin in having weaker interactions between the cytoplasmic ends of transmembrane (TM)3 and TM6, involving the conserved E/DRY sequences. These differences may be responsible for the relatively high basal activity and structural instability of the beta2AR, and contribute to the challenges in obtaining diffraction-quality crystals of non-rhodopsin GPCRs.