1H, 15N, 13C resonance assignment of folded and 8 M urea-denatured state of SUMO from Drosophila melanogaster

SUMO, an important post-translational modifier of variety of substrate proteins, regulates different cellular functions. Here, we report the NMR resonance assignment of the folded and 8 M urea-denatured state of SUMO from Drosophila melanogaster (dsmt3).     

Reversible monomer-oligomer transition in human prion protein

The structure and the dissociation reaction of oligomers PrP^oligo from reduced human prion huPrP^C(23–231) have been studied by ¹H-NMR and tryptophan fluorescence spectroscopy at varying pressure, along with circular dichroism and atomic force microscopy. The ¹H-NMR and fluorescence spectral feature of the oligomer is consistent with the notion that the N-terminal residues including all seven Trp residues, are free and mobile, while residues 105∼210, comprising the AGAAAAGA motif and S1-Loop-HelixA-Loop-S2-Loop-HelixC, are engaged in intra- and/or inter-molecular interactions. By increasing pressure to 200 MPa, the oligomers tend to dissociate into monomers which may be identified with PrP^C*, a rare metastable form of PrP^C stabilized at high pressure (Kachel et al., BMC Struct Biol 6:16). The results strongly suggest that the oligomeric form PrP^oligo is in dynamic equilibrium with the monomeric forms via PrP^C*, namely huPrP^C ⇆ huPrP^C* ⇆ huPrP^oligo.Key words: human prion, oligomer structure, pressure dissociation, reversible monomer-oligomer transition, circular dichroism, high pressure NMR, atomic force microscopy     

Dissecting the functional role of polyketide synthases in Dictyostelium discoideum: biosynthesis of the differentiation regulating factor 4-methyl-5-pentylbenzene-1,3-diol

Dictyostelium discoideum exhibits the largest repository of polyketide synthase (PKS) proteins of all known genomes. However, the functional relevance of these proteins in the biology of this organism remains largely obscure. On the basis of computational, biochemical, and gene expression studies, we propose that the multifunctional Dictyostelium PKS (DiPKS) protein DiPKS1 could be involved in the biosynthesis of the differentiation regulating factor 4-methyl-5-pentylbenzene-1,3-diol (MPBD). Our cell-free reconstitution studies of a novel acyl carrier protein Type III PKS didomain from DiPKS1 revealed a crucial role of protein-protein interactions in determining the final biosynthetic product. Whereas the Type III PKS domain by itself primarily produces acyl pyrones, the presence of the interacting acyl carrier protein domain modulates the catalytic activity to produce the alkyl resorcinol scaffold of MPBD. Furthermore, we have characterized an O-methyltransferase (OMT12) from Dictyostelium with the capability to modify this resorcinol ring to synthesize a variant of MPBD. We propose that such a modification in vivo could in fact provide subtle variations in biological function and specificity. In addition, we have performed systematic computational analysis of 45 multidomain PKSs, which revealed several unique features in DiPKS proteins. Our studies provide a new perspective in understanding mechanisms by which metabolic diversity could be generated by combining existing functional scaffolds.     

Biosynthesis of rubradirin as an ansamycin antibiotic from Streptomyces achromogenes var. rubradiris NRRL3061

The four overlapping cosmids from the rubradirin producer, Streptomyces achromogenes var rubradiris NRRL 3061, have 58 ORFs within a 105.6 kb fragment. These ORFs harbored essential genes responsible for the formation and attachment of four distinct moieties, along with the genes associated with regulatory, resistance, and transport functions. The PKS (rubA) and glycosyltransferase (rubG2) genes were disrupted in order to demonstrate a complete elimination of rubradirin production. The rubradirin biosynthetic pathway was proposed based on the putative functions of the gene products, the functional identification of sugar genes, and the mutant strains. The GeneBank accession number for the sequence reported in this paper is AJ871581.     

Synthesis and antibacterial activity of the C-7 side chain of 3-aminoquinazolinediones

A novel series of bacterial topoisomerase (3-aminoquinazolinediones) inhibitors are described. The side-chain SAR against Gram-positive and Gram-negative organisms as well as DNA gyrase activity is reported.     

Enteric neuroblasts require the phosphatidylinositol 3‐kinase pathway for GDNF‐stimulated proliferation

The enteric nervous system (ENS) develops from neural crest cells that enter the gut, migrate, proliferate, and differentiate into neurons and glia. The growth factor glial‐derived neurotrophic factor (GDNF) stimulates the proliferation and survival of enteric crest‐derived cells. We investigated the intracellular signaling pathways activated by GDNF and their involvement in proliferation. We found that GDNF stimulates the phosphorylation of both the PI 3‐kinase downstream substrate Akt and the MAP kinase substrate ERK in cultures of immunoaffinity‐purified embryonic avian enteric crest‐derived cells. The selective PI 3‐kinase inhibitor LY‐294002 blocked GDNF‐stimulated Akt phosphorylation in purified crest cells, and reduced proliferation in cultures of dissociated quail gut. The ERK kinase (MEK) inhibitors PD 98059 and UO126 did not reduce GDNF‐stimulated proliferation, although PD 98059 blocked GDNF‐stimulated phosphorylation of ERK. We conclude that the PI 3‐kinase pathway is necessary for the GDNF‐stimulated proliferation of enteric neuroblasts. © 2001 John Wiley & Sons, Inc. J Neurobiol 47: 306–317, 2001     

A comparative developmental pattern of enzymes of carbon metabolism and pentose phosphate pathway in mungbean and lentil nodules

The activities of enzymes of pentose phosphate pathway (PPP) viz. glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase and carbon metabolism viz. phosphoenol pyruvate carboxylase, NADP- isocitrate dehydrogenase and NADP-malic enzyme were measured in the plant and bacteroid fractions of mungbean (ureide exporter) and lentil (amide exporter) nodules along with the developing roots for comparison. The enzymes of pentose phosphate pathway in legume cytosol had higher activities at a stage of maximum nitrogenase activity and higher sucrose metabolism. However, bacteroids had only limited capacity for this pathway. The specific activities of these enzymes were greater in ureide than in amide exporter. CO₂ fixation via higher activity of phosphoenolpyruvate carboxylase in the plant part of the nodules in lentil might have been due to the greater synthesis of four carbon amino acids for amide export. The peak of NADP-isocitrate dehydrogenase in both legumes coincided with the pentose phosphate pathway enzymes at the time of high rates of sucrose metabolism and nitrogen fixation. Higher activities of NADP-malic enzyme were obtained in mungbean than in the lentil nodules. These findings are consistent with the role of these enzymes in providing reductant (NADPH) and substrates for energy yielding metabolism of bacteroids and carbon skeletons for ammonia assimilation.     

Microanatomy and cytochemistry of the gastro-respiratory tract of an air-breathing cobitidid fish,Lepidocephalichthys guntea

Gastro-respiratory tract of the loach,Lepidocephalichthys guntea has been studied with special reference to the nature of its mucus secreting epithelia. The mucous cells are strongly PAS-positive and their number per unit area (mm²) in the mucosal layers of oesophagus, intestinal bulb, intestine and rectum are 733, 531, 223 and 540, respectively. The air-breathing segment of the gut is completely devoid of neutral mucosubstances, and there is a predominance of acidic mucosubstances over the neutral ones throughout the digestive tube. The air-blood pathway of the accessory respiratory organ is about 2.6 μm which is higher than the values of air-breathing organs of other fishes.     

The Unusual Transmembrane Partition of the Hexameric Channel of the Hepatitis C Virus

1. Download high-res image (235KB)
2. Download full-size image

     

OCTAL: Optimal Completion of gene trees in polynomial time

Background

For a combination of reasons (including data generation protocols, approaches to taxon and gene sampling, and gene birth and loss), estimated gene trees are often incomplete, meaning that they do not contain all of the species of interest. As incomplete gene trees can impact downstream analyses, accurate completion of gene trees is desirable.

Results

We introduce the Optimal Tree Completion problem, a general optimization problem that involves completing an unrooted binary tree (i.e., adding missing leaves) so as to minimize its distance from a reference tree on a superset of the leaves. We present OCTAL, an algorithm that finds an optimal solution to this problem when the distance between trees is defined using the Robinson–Foulds (RF) distance, and we prove that OCTAL runs in \(O(n^2)\) time, where n is the total number of species. We report on a simulation study in which gene trees can differ from the species tree due to incomplete lineage sorting, and estimated gene trees are completed using OCTAL with a reference tree based on a species tree estimated from the multi-locus dataset. OCTAL produces completed gene trees that are closer to the true gene trees than an existing heuristic approach in ASTRAL-II, but the accuracy of a completed gene tree computed by OCTAL depends on how topologically similar the reference tree (typically an estimated species tree) is to the true gene tree.

Conclusions

OCTAL is a useful technique for adding missing taxa to incomplete gene trees and provides good accuracy under a wide range of model conditions. However, results show that OCTAL’s accuracy can be reduced when incomplete lineage sorting is high, as the reference tree can be far from the true gene tree. Hence, this study suggests that OCTAL would benefit from using other types of reference trees instead of species trees when there are large topological distances between true gene trees and species trees.