The recent origins of spliceosomal introns revisited

Does the intron/exon structure of eukaryotic genes belie their ancient assembly by exon-shuffling or have introns been inserted into preformed genes during eukaryotic evolution? These are the central questions in the ongoing ‘introns-early’ versus ‘introns-late’ controversy. The phylogenetic distribution of spliceosomal introns continues to strongly favor the intronslate theory. The introns-early theory, however, has claimed support from intron phase and protein structure correlations.     

Remembrance of things past: maintaining gene expression patterns with altered chromatin

Eukaryotic organisms have evolved mechanisms to stably preserve the gene expression patterns that determine cell fate. Recent advances have been made in understanding the DNA sequences and protein factors required to propagate gene activation or silencing. These studies suggest that, after gene activity states are selected during development, maintenance protein complexes provide a molecular memory of those states by altering a local domain of chromatin structure.     

Carbamoyl phosphate synthetase: a tunnel runs through it

The direct transfer of metabolites from one protein to another in a biochemical pathway or between one active site and another within a single enzyme has been described as substrate channeling. The first structural visualization of such a phenomenon was provided by the X-ray crystallographic analysis of tryptophan synthase, in which a tunnel of approximately 25 Å in length was observed. The recently determined three-dimensional structure of carbamoyl phosphate synthetase sets a new long distance record in that the three active sites are separated by nearly 100 Å.     

Two-component signal transduction systems in eukaryotic microorganisms

Conserved signal transduction pathways that use phosphorelay from histidine kinases through an intermediate transfer protein (H2) to response regulators have been found in a variety of eukaryotic microorganisms. Several of these pathways are linked to mitogen-activated protein kinase cascades. These networks control different physiological responses including osmoregulation, cAMP levels and cellular morphogenesis.     

Tubulin structure: insights into microtubule properties and functions

The structure of tubulin has recently been determined by electron crystallography, paving the way for a clearer understandin of the unique properties of tubulin that allow its varied functions within the cell. Some of the ongoing work on tubulin can be interpreted in terms of its structure, which can serve to guide future studies.     

Polarity and division site specification in yeast

A significant component of polarization in budding yeast involves the regulated restructuring of the actin cytoskeleton in response to defined cellular signals. Recent evidence suggests that such cytoskeletal organization arises through the action of large protein complexes that form in response to signals from small GTP-binding proteins, such as Cdc42, Rho, and Ras. These actin-organizing complexes may be fairly diverse, but generally consist of one or more central scaffold proteins, such as those of the formin class, that bind to signaling molecules and recruit actin-binding proteins to bring about desired polarizing events.     

Structure and dynamics of green fluorescent protein

Many marine organisms are luminescent. The proteins that produce the light include a primary light producer (aequorin or luciferase) and often a secondary photoprotein that red shifts the light for better penetration in the ocean. Green fluorescent protein is one such secondary protein. It is remarkable in that it autocatalyzes the formation of its own fluorophore and thus can be expressed in variety of organisms in its fluorescent form. The recent determination of its 3D structure and other physical characterizations are revealing its molecular mechanism of action     

Selection of biologically active peptides by phage display of random peptide libraries

Random peptide libraries displayed on phage are used as a source of peptides for epitope mapping, for the identification of critical amino acids responsible for protein—protein interactions and as leads for the discovery of new therapeutics. Efficient and simple procedures have been devised to select peptides binding to purified proteins, to monoclonal and polyclonal antibodies and to cell surfaces in vivo and in vitro.     

New structures of allosteric proteins revealing remarkable conformational changes

New three-dimensional structures of allosteric proteins reveal they have a flexible architecture that is instrumental to the regulation of protein function. Highlights are the structures of GroEL, pyruvate kinase, -3-phosphoglycerate dehydrogenase and the acetylcholine receptor. Furthermore, significant progress in understanding the nature of the intermediates involved in an allosteric reaction has been achieved through recent spectroscopic and crystallographic studies on haemoglobin.     

Synthesis of novel proteins

De novo and rational protein design are progressing towards the chemical synthesis of proteins with pre-selected structure and function. The data illustrate diverse experimental and computational approaches which test our comprehension of protein structure, hydrophobic core packing and global stability, especially of coiled-coil proteins. The incorporation of biologial cofactors, including hemes, as well as active sites, such as tht of iron superoxide dismutase, into designed proteins provides an exciting next step towards the synthesis of proteins with enzymatic function.     

Synapse maturation and structural plasticity at Drosophila neuromuscular junctions

The Drosophila larval neuromuscular junction has recently emerged as a powerful model system to characterize the cellular and molecular events involved in the formation and flexibility of synapses. The combination of molecular, genetic, electrophysiological and anatomical approaches has revealed, for example, the functional significance of the discs-large gene product (a novel synapse-organizing protein) in the nervous system. This protein is involved in the clustering of at least one ion channel and in the structural modification of glutamatergic synapses during target muscle growth. The manipulation of the genes encoding ion channels, components of second-messenger cascades, and cell adhesion molecules is beginning to tease apart the mechanisms underlying structural synaptic plasticity.     

Will combinatorial chemistry deliver real medicines?

Over the next decade, the impact of library synthesis will play a major role in shortening the lead optimization phase of drug discovery. The prognosis for combinatorial chemistry to discover fundamentally different new classes of therapeutically active small molecules against some of the more difficult biological targets is less certain. Expectations are high because the technology potentially allows us to sample available drug space by synthesizing all possible small molecule ligands (variously estimated to be between 10³⁰–10⁵⁰ compounds). Some caution is advised, however, since, despite recent increases in high-throughput screening of substantially greater numbers of synthetic compounds and natural products, we are not routinely finding a plethora of new structures. The outcome may be that combinatorial chemistry offers us the ability to work faster on finding ligands for well-established tractable targets, such as G-protein-coupled receptors, ion channels or proteases, rather than, say, the more complex protein—protein interactions which from the majority of targets in signal transduction pathways.     

Coordinate control of secondary metabolite production and asexual sporulation in Aspergillus nidulans

Microbial secondary metabolite production is frequently associated with developmental processes such as sporulation, but there are few cases where this correlation is understood. Recent work with the filamentous fungus Aspergillus nidulans has provided new insights into the mechanisms coordinating production of the toxic secondary metabolite sterigmatocystin with asexual sporulation. These processes have been shown to be linked through a common need to inactivate a heterotrimeric G protein dependent signaling pathway that, when active, serves to stimulate growth while blocking both sporulation and sterigmatocystin biosynthesis.     

Insights into the mechanisms of homologous recombination from the structure of RuvA

The recent structure determination of RuvA has provided the first insights into the structural basis for its interaction with Holliday junction DNA. Multiple copies of a helix-hairpin-helix motif which line the four grooves between the monomers in the tetrameric structure are thought to be involved in the interaction of the protein with its DNA target. This suggests that the four arms of the junction are held by RuvA in a fourfold symmetric arrangement and has fuelled ideas on the way in which components of the Ruv complex combine to catalyse the process of homologous recombination     

Unraveling the structures and modes of action of bacterial toxins

The mechanism by which a soluble protein converts into a protein that spans a membrane remains a central question in understanding the molecular mechanism of toxicity of bacterial protein toxins. Using crystallographic structures of soluble toxins as templates, the past year has seen a number of experiments that are designed to probe the membrane state using other structural methods. In addition, crystallographic information concerning the clostridial neurotoxins has emerged, suggesting a novel mechanism of pore formation and new relationships between toxin binding domains.     

Novel metal sites in protein structures

Recently, the three-dimensional structures of several novel metalloenzymes have been solved. Of special interest are those containing uncommon and/or not well characterized metals such as molybdenum, tungsten, nickel, vanadium and cobalt. Modulated by the protein environment, the specific properties of these metals and of special metal-binding cofactors such as siroheme and topa quinone are used to catalyze a vast array of fascinating reactions.     

Lipid polymorphism and biomembrane function

In recent years, several major developments have taken place in the biology, physical chemistry and technology of polymorphism of membrane lipids. These include the identification of polymorphic regulation of membrane lipid composition in Escherichia coli, the importance of nonbilayer lipids for protein functioning, the special packing properties of bilayers containing these lipids, and the crystalization of a membrane protein out of three dimensional bilayer networks (lipid cubic phases). These exciting developments bring us closer to understanding the paradox of the lipid bilayer structure of biomembranes and the molecular basis of membrane protein structure and function.     

Protein farnesyltransferase

In the past year, the crystal structure of αβ heterodimeric protein farnesyltransferase from rat was reported to a resolution of 2.25 Å. Farnesyltransferase catalyzes the essential post-transduction proteins. The structure provides a foundation for understanding the specificity and mechanism of protein prenylation and may aid in the design of new anticancer therapeutics.