Volume 18 Issue 5, May 2011

The CRISPR system is a prokaryotic immune system that depends on the Cascade protein complex and small guide RNAs (crRNAs). The Cascade complex, loaded with crRNA, is now characterized, the overall architecture of Cascade deduced, and the complex found to identify targets by formation of an R loop between the crRNA and dsDNA.

Hsp90 is a molecular chaperone with a wide array of client proteins, including the tumor suppressor p53. Now the conformation of p53 DNA-binding domain in the presence of full-length or fragments of Hsp90 is studied by HDX and NMR analyses. The results indicate that p53 adopts a looser, molten-globule-like state in the presence of Hsp90.

A central step in synaptic vesicle priming is the transition of syntaxin-1 between two very stable complexes: with Munc18 and with the other SNARE complex components. How this transition is promoted is now explored by NMR and FRET studies, revealing that the MUN domain of Munc13 uses weak interactions to pry open the closed syntaxin-1 in complex with Munc18.

Various models have been proposed for control of autoinhibition of the signaling adaptor protein Crk-II. A thermodynamic, kinetic and structural analysis reveals a crucial role for selective domain destabilization in tuning the responsiveness of Crk-II to activation.

A comprehensive HDX analysis of the vitamin D receptor–retinoid X receptor (VDR-RXR) reveals extensive allosteric communication within the same polypeptide where DNA or ligand can induce alterations in distant domains of the receptors. The work also shows communication between subunits in the heterodimer.

A combination of SAXS, SANS and FRET is used to explore the solution structures of three different nuclear receptor heterodimeric complexes (RXR with RAR, PPARγ or VDR), bound to different target DNA sequences. The work reveals the asymmetric shape of the complexes and the binding of a single coactivator molecule to RXR in each heterodimer.

IgE mediates allergic hypersensitivity and it is found tightly associated with its cellular receptor FcɛRI. Now crystal structure and binding studies reveal that the Cɛ2 domain from IgE Fc, which does not contact the receptor, contributes nevertheless to the high affinity by rendering the Cɛ3 domain more ordered, and thus minimizing the entropy gain upon receptor dissociation.

Spatial oscillation of the Min proteins directs septum formation during E. coli cell division. Using confocal and single molecule microscopy, the coordination of the membrane binding events that underlie this organization are now examined, giving insight into the dynamic events underlying this process.

The mechanism dictating which Raf isoform activates the MAPK pathway in melanoma has been unclear. This study now shows cross-talk between the MAPK and cAMP pathways, with B-Raf kinase specifically inhibited through interaction with Ras through a negative feedback loop in melanoma cells. C-Raf is activated through cAMP pathway inhibition by overexpression of PDE4 isoforms. This explains Raf isoform switching and offers a potential new therapeutic approach for melanoma.

Using phylogenetic analysis, ancestral forms of thioredoxin reflecting the sequences likely to have existed at key evolutionary points have now been synthesized and analyzed. Single-molecule analysis indicates that all of the reconstructed enzymes use the extant reaction mechanism but that the most ancient are optimally active at low pH and resistant to higher temperatures, conditions that would have existed on early earth.

Bacteriophages deliver their genome into the host cell through large proteinaceous assemblies called portal proteins. The first high-resolution structure of a portal protein from the Podovirae family reveals a ~1.1-MDa assembly formed by 24 proteins and a second structure uncovers an unexpected barrel-shaped domain that is essential for infectivity.

Endoplasmic reticulum aminopeptidase 1 (ERAP1) has a crucial role in antigen presentation because it trims peptide ligands to 8–10 residues so that they fit into the peptide-binding groove of MHC class I molecules. The structure of the ERAP1 in complex with bestatin, an aminopeptidase inhibitor, reveals how the enzyme´s catalytic center how the enzyme can preferentially process peptides 10–15 residues long while sparing shorter ones and provides the first evidence that substrate binding induces conformational changes.

A cryo-EM structure of the bacterial ribosome–SecYEG complex in a so-called Nanodisc allows for the molecular interpretation of the SecYEG complex in its natural lipid bilayer environment. Molecular dynamics simulations based on the structure reveal stable interactions between ribosomal RNA and the membrane that may contribute to the insertase activity of the protein-conducting channel.

Binding of the archaeal protein PbaA to proteasome precursors reveals the requirement for a C-terminal HbYX motif. This motif is also necessary for the function of the related yeast 20S proteasome assembly factor Pba1-Pba2. In addition, PbaA binds to mature 20S proteasomes if the β-subunit active sites are bound by inhibitors, implying an allosteric mechanism.

An intein controlled by redox in bacteria is created by engineering a disulfide bond between the catalytic cysteine residue and the flanking polypeptide. A natural analog of this arrangement from Pyrococcus abyssi is found to respond to redox conditions in Escherichia coli, suggesting that inteins may function as biological sensors.