FAST (Flexible Analysis by Software Tool) and CHAMP (CHemico-physical AMinoacidic Parameter data bank): a new tool to investigate protein structure

A flexible package designed to study protein structure is described.The package is devoted to the analysis of protein sequencesby drawing structural profiles of specific structure-relatedamino acid parameters. An Aminoacidic Parameters Data Bank (CHAMP)containing 32 different series of physico-chemical parametersof amino acids is available. Sequences can be loaded from anyASCII format data bank or from keyboard. The program possessesa routine which enables easy updating of the protein data bankand CHAMP Data Bank. FAST reads statistical correlations betweentwo plots in order to identify structural similarities. Plotscan be printed, saved or used for correlation, comparison orgraph overlap by using common spreadsheets (e.g. Lotus 123).Plots can be smoothed by a running mean or a running median.The program also has a special feature—a global flexibilityanalysis of proteins. The package runs on IBM or compatiblesand requires DOS 3.0 or later. Received on June 20, 1989; accepted on August 2, 1989     

Effect of Growth Stage and Initial Inoculum Level on Leaf Rust Development and Yield Loss Caused by Puccinia recondita f. sp. tritici

The effects of the combinations of leaf rust inoculation at different growth stages and initial inoculum levels on leaf rust development and yield of winter wheat cultivars, McNair 1003 and Coker 762 were evaluated. Disease onset stage and initial inoculum level affected the rate of leaf rust development and shape of the disease progress curves in both cultivars. Epidemics with common onset stages and different initial inoculum levels differed in area under the disease progress curve (AUDPC). Leaf rust epidemics initiated at Feeke's growth stages 5, 7, and 10 reduced yield in both cultivars. Leaf rust epidemics initiated early with high inoculum levels had the greatest deleterious effect on yield. Maximum losses due to leaf rust were 30–40 %. Yield loss was directly related to AUDPC when the AUDPC varied from 500 to 1700 in McNair 1003 and from 250 to 1700 in Coker 762. Yield reduction was mainly due to reduction in grain weight.     

Transformation of T lymphocytes by the v-fos oncogene

Activation of T lymphocytes through the T cell antigen receptor has been shown to stimulate a rapid and transient accumulation of c-fos mRNA and protein. Transfection of a normal murine T lymphocyte clone with the FBJ-v-fos oncogene resulted in generation of a cell line that was morphologically transformed, had lost the requirement for IL-2 for proliferation, and was tumorigenic in adult syngeneic mice; however, the transformed cells retained the ability to proliferate in response to IL-2. The transformed cells did not show constitutive expression of IL-2 or c-fos mRNA, although the promoter regions of both IL-2 and c-fos genes contain AP-1 sites that are expected to be targets for binding of Fos/Jun complexes. In contrast, the transformed T cells showed increased constitutive expression of IL-2R alpha and c-myc mRNA; these genes may represent cellular targets for transformation by v-fos and physiologic activation by c-fos. We discuss the possibility that these transformed cells behave as cells partially activated through the TCR, and that transformation occurs through a mechanism independent of IL-2.     

Cardiovascular risk factors in a French-Canadian population: resolution of genetic and familial environmental effects on blood pressure by using extensive information on environmental correlates.

Genetic and environmental influences on systolic blood pressure (SBP), diastolic blood pressure (DBP), and mean arterial blood pressure (MBP) were examined in 371 French-Canadian families by using path analysis. Familial environment was estimated with environmental indices constructed from as many as 14 (of a pool of more than 100) correlates of blood pressure (BP). Approximately 20% of the variance in BP can be accounted for by the composite index, and the types of variables and the direction of their effects vary as a function of age and of the multivariate context. Path analysis of the family data suggests that genetic heritability is relatively high in children (from 0.49 for SBP to 0.56 for MBP) but much smaller in adults (from 0.08 for DBP to 0.18 for SBP). The proportion of variability explained by familial environment is estimated to be the same in children and adults and is much higher than reported to date (from 0.30 for SBP to 0.42 for DBP). In addition, sibships share significant nontransmitted environmental effects, and there is no evidence to suggest specific maternal effects in the aggregation of BP. Two unique findings emerge from this study. First, unlike in most earlier studies, we were able to arrive at the same parsimonious model for each of the BP variables. Second, the familial environment accounts for a substantial proportion of the variability in BP, which has been considerably underestimated in earlier studies.     

Calcium ion binding to delta- and to beta-crystallins. The presence of the "EF-hand" motif in delta-crystallin that aids in calcium ion binding

Abnormal levels of endogenous calcium ions are known to induce eye lens opacity, and a variety of causative factors has been proposed, including calcium-mediated aggregation and precipitation of the lens proteins crystallins. We have specifically looked in some detail at the interaction of Ca2+ with various crystallins and its consequences. Lenses incubated in solutions containing 10 mM Ca2+ or 5 mM Tb3+ opacified. Fluorescence titration of crystallins with TbCl3 revealed that this ion binds to delta- and beta-crystallins in solution. Equilibrium dialysis showed that four Ca2+ ions bind to one delta-crystallin tetramer with an affinity of 4.3 x 10(3) M-1. Analysis of the amino acid sequence of delta-crystallin reveals the presence of a calmodulin-type "helix-loop-helix" or "EF-hand" calcium ion binding conformational motif in the region comprising residues 300-350. This is a novel feature of the molecule not reported so far. No other crystallins appear to have this motif. beta-Crystallin also binds four Ca2+ ions/aggregate unit of mass 160 kDa, with an affinity of 2.6 x 10(3) M-1, presumably in the midregion of the molecule that is rich in anionic and polar residues. Circular dichroism spectroscopy shows that the binding of calcium ion leads to subtle conformational changes in the molecules, notably in the tertiary structure.     

A new peptide in the FMRFamide family isolated from the CNS of the hawkmoth, Manduca sexta

We have purified a FMRFamide-like peptide from extracts of brain-subesophageal ganglion of the moth, Manduca sexta. The purification was monitored with a new, competitive ELISA, and accomplished with ion exchange and reverse-phase HPLC. The peptide structure was determined by a combination of tandem mass spectrometry and automated Edman degradation. The amino acid sequence of the peptide is less than Glu-Asp-Val-Val-His-Ser-Phe-Leu-Arg-Phe-amide (pEDVVHSFLRF-NH2). In a separate purification, an identical peptide was isolated from extracts of brain-associated neurohemal structures. We have named this peptide ManducaFLRFamide, to indicate its homology with other members of the "FMRFamide" family. In bioassays, chemically synthesized peptide increased the force of neurally evoked contractions in the major power-producing flight muscles, the dorsal longitudinal muscles. This observation suggests that hormonally released ManducaFLRFamide may play a role in sustaining or promoting the flight behavior necessary for mate-seeking (in males) or oviposition (in females) in sphingid moths.     

Analysis of human chromosome 21: correlation of physical and cytogenetic maps; gene and CpG island distributions.

Human chromosome 21 has been analyzed by pulsed-field gel electrophoresis using somatic cell hybrids containing limited regions of the chromosome and greater than 60 unique sequence probes. Thirty-three independent NotI fragments have been identified, totalling 43 million bp. This must account for essentially the entire long arm, and therefore gaps remaining in the map must be small. The extent of the pulsed-field map has allowed the direct correlation of the physical map with the cytogenetic map: translocation breakpoints can be unambiguously positioned along the long arm and the distances between them measured in base pairs. Three breakpoints have been identified, providing physical confirmation of cytogenetic landmarks. Information on sequence organization has been obtained: (i) 60% of the unique sequence probes are located within 11 physical linkage groups which can be contained in only 20% of the long arm; (ii) 9/21 genes are clustered within 4%; (iii) translocation breakpoints appear to occur within CpG island regions, making their identification difficult by pulsed-field techniques. This analysis contributes to the human genome mapping effort, and provides information to guide the rapid investigation of the biology of chromosome 21.     

Effect of Glutathione and N-Acetylcysteine on in Vitro and in VIVO Cardiac Toxicity of Doxorubicin

The effects of two sulfhydryl compounds, glutathione (GSH) and N-acetylcysteine (NAC), on the cardiotoxicity of doxorubicin (DXR) were tested on in vitro and in vivo models. DXR was administered to rats as 4 weekly i.v. doses of 3mg/kg. GSH (1.5 mmoles/kg), given i.v. 10 min before and 1 hr after DXR, was found to prevent the development of the delayed cardiotoxic effects of DXR, as assessed by electrocardiographic and mechanical parameters, as well as by histological examination of left ventricular preparations. In contrast, equimolar oral doses of NAC (1 hr before and 2hrs after DXR) were found to be ineffective. Both GSH and NAC prevented the negative inotropic effect produced by DXR on isolated rat atria. A good correlation exists between the cardioprotective effects of the two agents and their ability to enhance the non-protein sulfhydryl group content of the myocardium. Differences observed in vivo between GSH and NAC might be accounted for by pharmacokinetic factors.     

Modification of enzyme activity in reversed micelles through clathrate hydrate formation.

The physical phenomenon of clathrate hydrate formation in protein-containing reversed micelles is described. Hydrate formation in reversed micelles is a method of adjusting the water to surfactant molar ratio, wo, which influences micellar size. Lipase and alpha-chymotrypsin encapsulated in large reversed micelles of high wo show significant enhancements in activity when the micelle size is reduced through hydrate formation. Alternate methods of micelle size adjustments also show enhancements in activity. The implications for improving the activity of such encapsulated enzymes recovered from fermentation media through phase transfer into reversed micelles are discussed.