Complete sequences of the rRNA genes of Drosophila melanogaster

In this, the first of three papers, we present the sequence of the ribosomal RNA (rRNA) genes of Drosophila melanogaster. The gene regions of D. melanogaster rDNA encode four individual rRNAs: 18S (1,995 nt), 5.8S (123 nt), 2S (30 nt), and 28S (3,945 nt). The ribosomal DNA (rDNA) repeat of D. melanogaster is AT rich (65.9% overall), with the spacers being particularly AT rich. Analysis of DNA simplicity reveals that, in contrast to the intergenic spacer (IGS) and the external transcribed spacer (ETS), most of the rRNA gene regions have been refractory to the action of slippage-like events, with the exception of the 28S rRNA gene expansion segments. It would seem that the 28S rRNA can accommodate the products of slippage-like events without loss of activity. In the following two papers we analyze the effects of sequence divergence on the evolution of (1) the 28S gene "expansion segments" and (2) the 28S and 18S rRNA secondary structures among eukaryotic species, respectively. Our detailed analyses reveal, in addition to unequal crossing-over, (1) the involvement of slippage and biased mutation in the evolution of the rDNA multigene family and (2) the molecular coevolution of both expansion segments and the nucleotides involved with compensatory changes required to maintain secondary structures of RNA.      

Catalytic properties and active-site structural features of immobilized horse liver alcohol dehydrogenase

Alcohol dehydrogenase from horse liver was immobilized by covalent attachment to CNBr-Sepharose and by adsorption to octyl-Sepharose CL-4B, a hydrophobic analog of Sepharose. In each case, rate constants for the binding and release of coenzyme and for the oxidation of substrates were measured based on the concentration of accessible active-site zinc atoms determined by titration with a paramagnetic inhibitor. All rate constants were substantially reduced upon immobilization; however, the rate constant of immobilized enzyme for ethanol oxidation was independent of the immobilization method, whereas the rate constant for cyclohexanol oxidation was lower for enzyme immobilized to octyl-Sepharose. Consequently, the substrate specificity of the two immobilized enzyme samples differed by an order of magnitude. Moreover, EPR spectroscopy studies and computer graphic analyses of spin labels occupying three defined regions of the active-site domain indicated that the active-site conformation adjacent to the catalytic zinc atom was similar in the two samples while the conformation slightly further from the zinc atom was different. This result may explain why the two immobilized enzyme preparations exhibited the same rate constant toward a small substrate (ethanol) yet different rate constants toward a larger substrate (cyclohexanol), whose rate constant is expected to be sensitive to a larger portion of the active site.     

Echocardiographic assessment and percutaneous closure of multiple atrial septal defects

Percutaneous coronary intervention can be associated with distal embolization of thrombotic material causing myocardial necrosis and infarction. We discuss the role of intravascular imaging to guide the use of a distal protection device by describing the outcome of a young woman presenting with non-ST elevation myocardial infarction. Coronary angiography demonstrated an isolated minor stenosis in the proximal left anterior descending coronary artery with slight haziness beyond the lesion. Intravascular ultrasound confirmed an extensive thrombus overlying a bulky atherosclerotic plaque. A distal filter wire was therefore successfully used to reduce the risk of distal embolization. The use of intravascular ultrasound in patients presenting with acute coronary syndrome may reveal large thrombi that are difficult to image using conventional angiographic techniques. Intravascular ultrasound can therefore be used as a tool to select lesions requiring distal protection.     

A systematic screen for genes expressed in definitive endoderm by Serial Analysis of Gene Expression (SAGE)

Background  

The embryonic definitive endoderm (DE) gives rise to organs of the gastrointestinal and respiratory tract including the liver, pancreas and epithelia of the lung and colon. Understanding how DE progenitor cells generate these tissues is critical to understanding the cause of visceral organ disorders and cancers, and will ultimately lead to novel therapies including tissue and organ regeneration. However, investigation into the molecular mechanisms of DE differentiation has been hindered by the lack of early DE-specific markers.     

Accurate prediction of protein secondary structure and solvent accessibility by consensus combiners of sequence and structure information

Background  

Structural properties of proteins such as secondary structure and solvent accessibility contribute to three-dimensional structure prediction, not only in the ab initio case but also when homology information to known structures is available. Structural properties are also routinely used in protein analysis even when homology is available, largely because homology modelling is lower throughput than, say, secondary structure prediction. Nonetheless, predictors of secondary structure and solvent accessibility are virtually always ab initio.     

Methods for peptide identification by spectral comparison

Background  

Tandem mass spectrometry followed by database search is currently the predominant technology for peptide sequencing in shotgun proteomics experiments. Most methods compare experimentally observed spectra to the theoretical spectra predicted from the sequences in protein databases. There is a growing interest, however, in comparing unknown experimental spectra to a library of previously identified spectra. This approach has the advantage of taking into account instrument-dependent factors and peptide-specific differences in fragmentation probabilities. It is also computationally more efficient for high-throughput proteomics studies.