SDSE: A software package to simulate the evolution of a pair of DNA sequences

An algorithm to simulate DNA sequence evolution under a generalstochastic model, including as particular cases all the previouslyused schemes of nucleotide substitution, is described. The simulationis carried out on finite, variable length, DNA sequences througha strict stochastic process, according to the particular substitutionrates imposed by each scheme. Five FORTRAN programs, runningon an IBM PC and compatibles, carry out all the tasks neededfor the simulation. They are menu driven and interfaced to thesystem through a principal menu. All sequence data files usedand generated by the SDSE package conform to the standard GenBankdatabase format, thus allowing the use of any sequence retrievedfrom this databank, as well as the application of other packagesto analyse, manipulate or retrieve simulated sequences. Received on August 23, 1988; accepted on November 15, 1988     

A 14-kilodalton selenium-binding protein in mouse liver is fatty acid-binding protein

In a previous study, we purified three selenium-binding proteins (molecular masses 56, 14, and 12 kDa) from mouse liver using column chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The aim of the present study was to determine the amino acid sequence of the 14-kDa protein thereby establishing any relationship with known proteins. Although the amino terminus of the 14-kDa protein was blocked, separate in situ digestions of the protein with endoproteinases Glu-c and Lys-c gave overlapping peptides that provided a continuous sequence of 93 amino acids. This sequence exhibited a 92.5% sequence homology with rat liver fatty acid-binding protein. In situ enzymatic digestion and partial sequencing of a 12-kDa selenium-binding protein revealed identical homology to the 14-kDa protein. The 14-kDa protein bound specifically to an oleate-affinity column from which the protein and 75Se coeluted. Delipidation or sodium dodecyl sulfate treatment failed to remove 75Se from the protein, indicating that the selenium moiety was tightly bound to the protein. These observations confirm that the mouse liver selenium-binding 14-kDa protein is a fatty acid-binding protein. The nature of the selenium linkage to the protein still needs to be explored.     

In search of mammary gland stem cells

Summary The evidence for mammary epithelial stem cells and their phenotypic characteristics in normal and neoplastic development is reviewed. The presence of stem cells in all parts of the mammary parenchyma at all stages of differentiation has been demonstrated by transplantation experiments. The phenotypic characterization of stem cells has been defined by a battery of monospecific antibodies. These studies suggest that a mammary epithelium stem cell compartment exists in the basal layer of the gland as well as in the end bud. Whether these same stem cells are expressed in mammary preneoplasias and neoplasias has not been answered conclusively. Phenotypic markers specific for stem cells and stably expressed in transformed cell populations are needed to follow the fate of stem cells.Dedicated to Professor Stuart Patton on the occassion of his 70th birthday.     

Neuron-specific enolase (NSE)-like and neurofilament protein (NFP)-like immunoreactivities in the rat dorsal root ganglia and sciatic nerve

The presence of neuron-specific enolase (NSF) and neurofilament proteins (NFP) immunoreactivities (IR) was investigated in dorsal root ganglia (DRG) of adult rats at cervical, thoracic, lumbar and sacral levels. All neurons display NSE-like IR with a variable intensity of immunostain which is not related to the neuronal size. Conversely, the antibody against all three proteic subunits of NFP no labelled the primary sensory neurons, whereas the intraganglionic axons and dorsal root of spinal nerves result positives. In the sciatic nerve the immunoreactivity was similar for NSE- and NFP-like IR. No regional differences were found among the different levels of DRG for NSE-like IR. The present results demonstrate heterogeneity in the neurons of the rat. DRG for NSE-like IR, and differences between sensory neurons and fibers in the distribution of NFP-like IR.     

Polymerization of τ into Filaments in the Presence of Heparin: The Minimal Sequence Required for τ - τ Interaction

Abstract: Paired helical filaments isolated from the brains of patients with Alzheimer's disease are composed of a major protein component, the microtubule-associated protein termed τ, together with other nonprotein components, including heparan, a glycosaminoglycan, the more extensively sulfated form of which is heparin. As some of these nonprotein components may modulate the assembly of τ into filamentous structures, we have analyzed the ability of the whole τ protein or some of its fragments to self-assemble in the presence of heparin. Different τ fragments, all of them containing some sequences of the tubulin-binding motif, can assemble in vitro into filaments. We have also found formation of polymers with the 18-residue-long peptide corresponding to the third tubulin-binding motif of τ. This suggests that the ability of τ for self-assembly could be localized in a short sequence of amino acids present in the tubulin-binding repeats of the τ molecule.     

Gram-scale purification of eicosapentaenoic acid (EPA, 20:5n-3) from wetPhaeodactylum tricornutum UTEX 640 biomass

Eicosapentaenoic acid (EPA, 20:5n-3) was obtained from the microalgaPhaeodactylum tricornutum following a three-step process: fatty acid extraction by direct saponification of wet biomass, polyunsaturated fatty acid (PUFA) concentration by formation of urea inclusion compounds and EPA isolation by preparative HPLC. Direct saponification of wet biomass was carried out with KOH-ethanol (96% v:v) (1 h, 60 °C), extracting 91% of the EPA. PUFAs were concentrated by the urea method with an urea/fatty acid ratio of 4:1 at a crystallization temperature of 28 °C using methanol as the urea solvent. An EPA concentration ratio of 1.5 (55.2/36.3) and recovery of 79% were obtained. This PUFA concentrate was used to obtain 95.8% pure EPA by preparative HPLC, using a reverse-phase column (C₁₈, 4.7 cm i.d. × 30 cm) and methanol-water (1% AcH) 80:20 w/w as the mobile phase. Ninety-seven per cent of EPA loaded was recovered and 70% EPA present in theP. tricornutum biomass was recovered in a highly pure form by means of this three-step downstream processing. In each of the HPLC preparative runs, 635 mg PUFA concentrate were loaded, obtaining 326 mg of a highly concentrated EPA fraction (2.46 g d^–1). Finally, a preliminary cost statement has been calculated.