The levels of ζ, γ, and δ chains in patients with Hb H disease

Summary Details are given of a study of blood samples from 24 patients with Hb H disease from different Mediterranean countries and from the Far East. Four different types of -thal-1 (--) were observed, namely-() ( 20.5-kb deletion);--MED-I ( 17.5-kb deletion);--MED-II (>26.5-kb deletion); and--SEA ( 18-kb deletion, in Orientals only). The -thal-2 was mainly of the deletion type (16 with the 3.7-kb deletion; 1 with the 4.2-kb deletion), while 4 of the 7 patients with a nondeletional type had the five-nucleotide deletion at the donor splice site of the first intron of the 2 gene. All patients had a mild-to-moderate hemolytic anemia; no significant differences in hematology were observed between the groups. Hb A₂ was decreased to about one-third of the normal level. The Hb H formation varied considerably and its quantitation was not always satisfactory. Patients with Hb H disease due to any -thal-1 combined with a nondeletional -thal-2 had the highest Hb H levels and a more marked anemia. The chain production was small and absent in patients with the MED-II type of -thal-1 because this deletion included the and genes. The highest chain levels were present in the four patients with the SEA type of -thal-1. The chain production was increased, particularly in patients with a mutation of C T at position-158 to the ^G globin gene. This chain was primarily present as Hb Bart's (or ₄) and only about 15% was recovered as Hb F or ₂₂. The evaluation of the rate of chains produced in these patients was greatly facilitated by data from one patient who had Hb H disease and a heterozygosity for the ^A-⁺. The low levels of Hb A₂ and of Hb F (relative to Hb Bart's) can be explained by a decreased affinity of chains for and chains as compared with chains in conditions of severe chain deficiency.     

Dopamine-1-mediated stimulation of phospholipase C activity in rat renal cortical membranes

Phospholipase C (PL-C) mediates transduction of neurotransmitter signals across membranes via hydrolysis of phosphatidylinositol-4,5-bisphosphate (PIP2), leading to generation of second messengers inositol-1,4,5-trisphosphate and diacylglycerol. In this study, dopamine-1 (DA-1) but not dopamine-2 (DA-2) agonists were shown to stimulate PL-C activity in renal cortical membranes. The DA-1 agonist, SKF 82526, stimulated the release of inositol phosphates from renal cortical membranes prelabeled with [3H]myoinositol. The majority of the label (75%) was found in phosphatidylinositol followed by PIP2 (15%) and phosphatidylinositol-4-phosphate (10%). A DA-1 specific effect on PL-C activity was also observed in an in vitro assay of PL-C activity in renal cortical membranes and basolateral and brush border membranes using [3H]PIP2 as the substrate. Dopamine and SKF 82526 stimulated the release of inositol phosphates from added [3H]PIP2 in a concentration-dependent manner. This release was blocked by the DA-1 antagonist SCH 23390 but not by the alpha-adrenergic antagonists phentolamine and prazosin. In contrast, the DA-2 agonist LY 171555 had no effect on inositol phosphate release. Guanosine 5'-(3-O-thio)triphosphate enhanced while guanyl-5'-yl thiophosphate attenuated the DA-1 agonist-stimulated PL-C activity. PL-C activity as measured by [3H]PIP2 hydrolysis had a pH optimum of 6.5, was inhibited by Mg2+ concentrations above 1 mM, was linear with time and protein concentration, and was sensitive to phosphatidylserine and calcium concentrations. We conclude that PL-C is activated by DA-1 but not DA-2 agonists in renal cortical membranes as well as both the basolateral and brush border renal tubular membranes. It is speculated that this action may mediate the natriuretic effects of dopamine in renal tubular epithelia.     

Unfolding of trp repressor studied using fluorescence spectroscopic techniques.

The unfolding properties of the trp repressor of Escherichia coli have been studied using a number of different time-resolved and steady-state fluorescence approaches. Denaturation by urea was monitored by the average fluorescence emission energy of the intrinsic tryptophan residues of the repressor. These data were consistent with a two-state transition from dimer to unfolded monomer with a free energy of unfolding of 19.2 kcal/mol. The frequency response profiles of the fluorescence emission brought to light subtle urea-induced modifications of the intrinsic tryptophan decay parameters both preceding and following the main unfolding transition. The increase of lifetime induced by urea required higher concentrations of urea than the increase in the total intensity described by Gittelman and Matthews [(1990) Biochemistry 29, 7011]. This indicates that the intensity increase has both dynamic and static origins. To assess the effect of tryptophan binding upon repressor stability, and to determine whether repressor oligomerization would be detectable in an unfolding experiment, we examined denaturation profiles of repressor labeled with the long-lived fluorescence probe 5-(dimethylamino)naphthalene-1-sulfonyl (DNS), by monitoring the average rotational correlation time of the probe. These experiments revealed a protein concentration dependent transition at low urea concentrations. This transition was promoted by tryptophan binding. We ascribe this transition to urea-induced dissociation of repressor tetramers. The main unfolding transition of the dimer to unfolded monomer was also observable using this technique, and the free energies associated with this transition were 18.3 kcal/mol in the absence of tryptophan and 24.1 kcal/mol in its presence, demonstrating that co-repressor binding stabilizes the repressor dimer against denaturation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Trinuclear iridium and rhodium complexes: the solution to a puzzle involving the multiple coordination possibilities of 1,8-naphthyridine-2-one ligands

The multiple coordination possibilities of 1,8-naphthyridine-2-one (HOnapy) and 5,7-dimethyl-1,8-napthyridine-2-one (HOMe₂napy) ligands allow the synthesis of a variety of tri- di- and mononuclear complexes, showing fluxional behaviour and frequent exchange of the coordinated ML₂ fragments. Thus, reactions of [M₂(μ-OMe)₂(cod)₂] (cod = 1,5-cyclooctadiene) with HOnapy and HOMe₂napy yield the compounds of the general formula [M(μ-OR₂napy) (cod)]_n (M = Ir, R = Me (1a, 1b, H (2); M = Rh, R = Me (3a, 3b). They crystallise as inconvertible yellow (a) and purple/orange (b) forms and also show a puzzling behaviour in solution. X-ray diffraction studies on both forms (3a, 3b) and spectroscopic data reveal that the yellow forms are mononuclear complexes whilst the dark-coloured crystals contain dinuclear complexes. In solution, the nuclearity of the complexes depends on the solvent. In addition both types of complexes are fluxional. The mixed-ligand complexes [M₂(μ-OMe₂napy)₂(CO)₂(cod)] M = Ir (5), Rh (6) have been isolated and characterised; they are found to be intermediates in the synthesis of the trinuclear complexes [M₃(μ₃-OMe₂napy)₂(CO)₂(cod)₂]⁺ M = Rh (8), Ir (9). Reactions of [IrCl(CO)₂(NH₂-p-tolyl] with the complexes [Rh(μ-OR₂napy)(diolefin)]_n followed by addition of a poor donor anion is a general one-pot synthesis for the hetertrinuclear complexes [Rh₂Ir(μ₃-OR₂napy)₂(CO)₂(diolefin)₂]⁺ (R=Me, DIOLEFIN = cod (10), tetrafluorobenzo-barrelene (tfbb) (11), 2,5-norbornadiene (nbd) (12); R=H, DIOLEFIN=cod (13)). This synthesis follows a stepwise mechanism from the mononuclear to the trinuclear complexes in which mixed-ligand heterodinuclear complexes are involved as intermediates of the type [(diolefin)Rh(μ-OMe₂napy)₂Ir(CO)₂]. Heteronuclear complexes which possess the core [RhIr₂]³⁺, such as [RhIr₂(μ₃-OR₂napy)₂(CO)₂(cod)₂]BF₄ (R=Me (14), H (15)), result from the reaction of 1 or 2 with [Rh(CO)₂S_x]⁺ (S = solvent). The trinuclear complexes undergo two chemically reversible one-electron oxidation processes. The chemical oxidation of 10, 14 and 9 with silver salts gives the mixed-valence trinuclear radicals [Rh₂Ir(μ₃-OMe₂napy)₂(CO)₂(cod)₂]²⁺ (16), [RhIr₂(μ₃-OMe₂napy)₂(CO)₂(cod)₂]²⁺ (17) and [Ir₃(μ₃-OMe₂napy)₂(CO)₂(cod)₂]²⁺ (18), which have been isolated as the perchlorate and tetrafluoroborate salts. The EPR spectrum of 16 indicates that the unpaired electron is essentially in an orbital delocalised on the metals. The molecular structures of the complexes 3a, 3b, 6, 10b and 16a are described. Crystals of 3a are triclinic, P-1, with a = 9.7393(2), b = 14.0148(4), c = 16.0607(4) Å, α = 88.122(3), β = 83.924(3), γ = 87.038(3)°, Z = 4; 3b crystallises in the Pna2_i orthorhhombic space group, with a = 16.7541(3), B = 11.7500(8), c = 17.7508(7) Å, Z = 4; complex 6 is packed in the monoclinic space group P2_i/c, a = 9.6371(1), b = 11.8054(4), c = 27.2010(9) Å, β = 90.556(4)°, Z = 4; crystals of 10b are monoclinic, P2₁/n, with a = 17.546(7), b = 13.232(6), c = 17.437(8) Å, β = 106.18(1)°, Z = 4; crystals of 16a are triclinic, P-1, with a = 10.318(4), b = 12.562(6), C = 19.308(8) Å, α = 92.12(8), β = 97.65(9), γ = 90.68(5)°, Z = 2. The five different structures show the coordination versatility of the OMe₂napy molecule as ligand, which behaves as a N,N′-chelating (3a), bidentate N,O-donor (3b, 6), or as a tridentate N,N′,O-donor bridging ligand (10b, 16a).     

Effect of distance,aggregation, and habitat on levels of seed predation for two mammal — dispersed neotropical rain forest tree species

The effect of seed aggregation and distance from conspecific trees on seed predation was experimentally examined for two neotropical tree species, Macoubea guianensis (Apocynaceae) and Pouteria sp. (Sapotaceae) in a lowland tropical rain forest in northeastern Peru. Results of these experiments are discussed in the context of the Janzen-Connell model (Janzen 1970; Connell 1971), which predicts decreased seed survival near parent trees due to either density-or distance-responsive mortality, and Howe's model (Howe 1989) which predicts that trees with seeds dispersed in clumps (aggregated) will not suffer density-dependent predation, and will have higher survival of seeds near the parent tree than other trees. We also examined whether predation on seeds of these species was affected by seed placement in or near 30-m-wide strips regenerating after clear-cutting. Both species appeared to be mammal-dispersed but differed in how frugivores handled seeds, seed size, overall fruit crop size, and gemination time. Neither of the two species studied appeared to suffer seed predation in a manner predicted by the Janzen-Connell model, and patterns of seed predation for only one of the species was similar to predictions of Howe's model. For neither species did seed predation along the edge of, or in the center of, regenerating clear cuts differ from predation 15 m into the primary forest. For Pouteria, seed predation in and near regnerating strips was significantly greater than around forest trees, but the opposite pattern held for Macoubea. Overall, seed predation was much greater on Macoubea. The difference in seed predation for these two species was most likely a result of differences in the types of seed predators that attacked these two species.     

Xp-duplications with and without sex reversal

Duplications in Xp including the DSS (dosage sensitive sex reversal) region cause male to female sex reversal. We investigated two patients from families with Xp duplications. The first case was one of two sisters with karyotype 46,XY, der(22), t(X;22)(p11.3;p11)mat and unambiguous female genitalia. The living sister was developmentally retarded, and showed multiple dysmorphic features and an acrocallosal syndrome. The second case was a boy with a maternally inherited direct duplication of Xp21.3-pter with the breakpoint close to the DSS locus. He had multiple abnormalities and micropenis, but otherwise unambiguous male genitalia. We performed quantitative Southern blot analysis with probes from Xp22.13 to p21.2 to define the duplicated region. Clinical, cytogenetic, and molecular data from both patients were compared with those of previously reported related cases. A comparison of the extragenital symptoms revealed no differences between patients with or without sex reversal. In both cases, the symptoms were non-specific. Among 22 patients with a duplication in Xp, nine had unambiguous female genitalia and a well-documented duplication of the DSS region. Two patients with duplication of DSS showed ambiguous external genitalia. From these data, we conclude that induction of testicular tissue may start in these patients, but that the type of genitalia depends on the degree of subsequent degeneration by a gene in DSS.     

Molecular-size corrections to the strength of the hydrophobic effect: a critical review

Large increases in the strength of the hydrophobic effect and, consequently, in the estimates of the hydrophobic contribution to the thermodynamics of protein folding (and other biologically-relevant processes), have been recently advocated on the basis of the application, to model transfer thermodynamic data, of corrections for the solute/solvent size disparity. In this work we first examine the effect of molecular-size corrections on the values calculated from several types of model transfer data. For the transfer of a solute from an organic solvent to water, the above increase is exclusively associated with the application of a solute/water molecular-size correction. Secondly, we critically review and assess the several theoretical arguments that lead to these corrections. In particular, we show that, contrary to previous claims in the literature, the analysis of dissolution processes in terms of ideal-gas, intermediate states does not lead to the molecular-size correction term, but only to expressions equivalent (although not strictly identical) to those derived from the well-known Ben-Naim's statistical-mechanical approach. In general, the several analyses offered or discussed in this work disfavor the application of the solute/water molecular-size corrections.     

Effect of abscisic acid, osmolarity and partial desiccation on the development of recalcitrant mango somatic embryos

Inhibition of mango somatic embryo growth was inducedin vitro by treatments for 4 or more weeks with abscisic acid (0–100 M ABA) with and without high osmolarity provided by mannitol (0–10%). High osmolarity and ABA significantly affected somatic embryo length, precocious germination and the production of good quality secondary somatic embryos. High osmolarity also affected root elongation. Abscisic acid was more effective in suppressing growth and development of 0.5 cm-length somatic embryos than smaller somatic embryos. Development beyond the heart stage was significantly inhibited by both ABA and mannitol treatments. The recovery of good quality somatic embryos was enhanced by high levels of ABA (100 M) with and without mannitol (0–5%). Somatic embryos that had been pulsed with ABA were partially desiccated at different relative humidities. Weight loss was affected only by relative humidity; and ABA did not enhance desiccation tolerance.Abbreviations ABA Abscisic acid - 2,4-D 2,4-Dichlorophenoxyacetic acid - MM1 Mango maturation medium - RH Relative humidity