Comparative studies of allozyme loci in Drosophila simulans and Drosophila melanogaster. I. Three dipeptidase loci

Genetic variation at three dipeptidase loci (Dip-A, Dip-B, and Dip-C) in Drosophila simulans was analyzed by starch gel electrophoresis. Dip-A was found to be polymorphic in four populations, while Dip-B and Dip-C were found to be polymorphic in one. The numbers of different alleles found at each respective locus were: Dip-A, two; Dip-B, two; and Dip-C, three. Dip-A was genetically mapped at 57.9 on the second chromosome, and Dip-B and Dip-C at 80.9 and 87.9 on the third chromosome, respectively. Neither Dip-B nor Dip-C has been mapped in D. melanogaster because both loci are apparently monomorphic. Their map positions in D. simulans with respect to flanking markers whose homologous genes have been cytogenetically localized in D. melanogaster suggested that they might be mapped cytogenetically by using available deficiencies in D. melanogaster. Accordingly, by the construction of interspecific hybrids which carried deficiencies of melanogaster and an allele of simulans with a mobility different from that of the fixed melanogaster allele, Dip-B and Dip-C were localized between 87F12-14 and 88C1-3 and between 87B5-6 and 87B8-10, respectively, in the salivary gland chromosomes of D. melanogaster. The similarity between these two species is discussed on the basis of these findings.     

Parameter Estimation for the Bateman Function via Moments of the Empirical Curve

The parameters of the function f(t)=c(e^?at-e^?bt) are related in a simple way to the moments ^∞ tⁿf(t)^dt(n=0, 1, 2). Using empirical values of f, the moments can be estimated by numerical integration. Therefrom estimates of the parameters are obtained by elementary algebra.     

Material Balance and Additional Effect of Second Organic Solutes in the Radiolysis of Aqueous Methionine Solution

The gamma-radiolysis of methionine in 10^?3 m aqueous solution was studied. From the yield vs. dose plots, the following values were obtained: G(?Methionine)=3.0, G(Methionine sulfoxide)=0.30, G(α- Aminobutyric acid)=0.46, G(3-Methylthio propylamine)=0.59, G(Methional)=0.08, G(Carboxylic acid)=0.24, G(Mercaptan + Disulfide)=0.6l, G(Ammonia)=1.48, and G(Carbon dioxide)=1.45. From these results, it was concluded that major reaction was deamination and decarboxylation by OH radical attack.

Addition of methanol as the second solute at concentration above 4 × 10^?3 m could not lower G(Methionine sulfoxide), suggesting the reaction of methionine with secondary radical produced from methanol.

Existence of similar reaction was suggested in the radiolysis of methionine in aqueous pork extract.     

Kinetic simulation studies on the transient formation of the oxo‐iron(IV) porphyrin radical cation during the reaction of iron(III) tetrakis‐5,10,15,20‐(N‐methyl‐4‐pyridyl)‐porphyrin with hydrogen peroxide in aqueous solution

High‐valent oxo‐iron(IV) species are commonly proposed as the key intermediates in the catalytic mechanisms of iron enzymes. Water‐soluble iron(III) tetrakis‐5,10,15,20‐(N‐methyl‐4‐pyridyl)porphyrin (Fe(III)TMPyP) has been used as a model of heme‐enzyme to catalyse the hydrogen peroxide (H₂O₂) oxidation of various organic compounds. However, the mechanism of the reaction of Fe(III)TMPyP with H₂O₂ has not been fully established. In this study, we have explored the kinetic simulation of the reaction of Fe(III)TMPyP with H₂O₂ and of the catalytic reactivity of FeTMPyP in the luminescent peroxidation of luminol. According to the mechanism that has been established in this work, Fe(III)TMPyP is oxidized by H₂O₂ to produce (TMPyP)^·+Fe(IV)=O (k1 = 4.5 × 10⁴/mol/L/s) as a precursor of TMPyPFe(IV)=O. The intermediate, (TMPyP)^·+Fe(IV)=O, represented nearly 2% of Fe(III)TMPyP but it does not accumulate in suf?cient concentration to be detected because its decay rate is too fast. Kinetic simulations showed that the proposed scheme is capable of reproducing the observed time courses of FeTMPyP in various oxidation states and the decay pro?les of the luminol chemiluminescence. It also shows that (TMPyP)^·+Fe(IV)=O is 100 times more reactive than TMPyPFe(IV)=O in most of the reactions. These two species are responsible for the initial sharp and the sustained luminol emissions, respectively. Copyright © 2003 John Wiley & Sons, Ltd.     

A model for the regulation of brain adenylate cyclase by ionic equilibria

Multiple-equilibrium equations were solved to investigate the individual and separate effects of Mg²⁺, Mn²⁺, Ca²⁺, ATP^4–, and their complexes on the kinetics of brain adenylate cyclase. The effects of divalent metals and/or ATP^4– (in excess of their participation in complex formation) were determined and, from the corresponding apparent affinity values, the following kinetic constants were obtained:K _m(MgATP)=1.0 mM,K _i(ATP^4–)=0.27 mM,K _m(MnATP)=0.07 mM, andK _i(CaATP)=0.015 mM. MgATP, MnATP, ATP^4–, and CaATP were shown to compete for the active site of the enzyme. Hence, it is proposed that endogenous metabolites with a strong ligand activity for divalent metals, such as citrate and some amino acids, become integrated into a metabolite feedback control of the enzyme through the release of ATP^4– from MgATP. Ca²⁺ fluxes may participate in the endogenous regulation of adenylate cyclase by modifying the level of CaATP. The free divalent metals show an order of affinityK _0.5(Ca²⁺)=0.02 mM,K _0.5(Mn²⁺)=3.8 mM,K _0.5(Mg²⁺)=4.7 mM, and an order of activity Mn²⁺>Mg²⁺>Ca²⁺. The data indicate that Mn²⁺ and Mg²⁺ ions may compete for a regulatory site distinct from the active site and increaseV _m without changingK _m(MgATP),K _m(MnATP), orK _i(ATP^4–). The interactions of ATP^4– and CaATP, which act as competitive inhibitors of the reaction of the enzyme with the substrates MgATP and MnATP, and Mg²⁺ and Mn²⁺, which act as activators of the enzyme in the absence of hormones, are shown to follow the random rapid equilibrium BiBi group-transfer mechanism of Cleland with the stipulation that neither Mg²⁺ nor Mn²⁺, in excess of their respective participation in substrate formation, are obligatorily required for basal activity. ATP^4– and CaATP are involved in dead-end inhibition. For MgCl₂ saturation curves at constant total ATP concentration, the computer-generated curves based on the RARE BiBi model predict a change in the Hill cooperativityh from a basal value of 2.6, when Mg²⁺ is not obligatorily required, to 4.0 when the addition of hormones or neurotransmitters induces an obligatory requirement for Mg²⁺.Abbreviations used: Me, divalent metal; Me_T (Mg_T or Mn_T), total Me (Me²⁺ and its complexes); ATP_T, total ATP (ATP^4– and its complexes).     

Determination of celiprolol and oxprenolol in human plasma by high-performance liquid chromatography and the analytical error function

Two reversed-phase HPLC methods with UV detection to quantify celiprolol and oxprenolol in human plasma are described. The analytical methods for the determination of both drugs used the same reversed-phase HPLC column, mobile phase and extraction procedure. Linearity was obtained in the ranges 15.63–1000 and 25–800 ng/ml for celiprolol and oxprenolol, respectively. Intra-day and inter-day variation was lower than 14%. After validation of the methods, analytical error functions were established as S.D. (ng/ml)=3.096+0.041C for celiprolol and S.D. (ng/ml)=8.906+8.075·10⁻⁸C³ for oxprenolol.     

Stereoselective and driving-force-dependent photoinduced electron-transfer reactions of zinc myoglobin with optically active N,N′-dimethylcinchoninium and N,N′-dimethylcinchonidinium ions

In order to understand the detailed mechanism of the stereoselective photoinduced electron-transfer (ET) reactions of zinc-substituted myoglobin (ZnMb) with optically active molecules by flash photolysis, we designed and prepared new optically active agents, such as N,N′-dimethylcinchoninium diiodide ([MCN]I₂) and N,N′-dimethylcinchonidinium diiodide ([MCD]I₂). The photoexcited triplet state of ZnMb, ³(ZnMb)*, was successfully quenched by [MCN]²⁺ and [MCD]²⁺ ions to form the radical pair of ZnMb cation (ZnMb^·+) and reduced [MCN]^·+ and [MCD]^·+, followed by a thermal back ET reaction to the ground state. The rate constants (k _q) for the ET quenching at 25 °C were obtained as k _q(MCN)=(1.9±0.1)×10⁶ M⁻¹ s⁻¹ and k _q(MCD)=(3.0±0.2)×10⁶ M⁻¹ s⁻¹, respectively. The ratio of k _q(MCD)/k _q(MCN)=1.6 indicates that the [MCD]²⁺ preferentially quenches ³(ZnMb)*. The second-order rate constants (k _b) for the thermal back ET reaction from [MCN]^·+ and [MCD]^·+ to ZnMb^·+ at 25 °C were k _b(MCN)=(0.79±0.04)×10⁸ M⁻¹ s⁻¹ and k _b(MCD)=(1.0±0.1)×10⁸ M⁻¹ s⁻¹, respectively, and the selectivity was k _q(MCD)/k _q(MCN)=1.3. Both quenching and thermal back ET reactions are controlled by the ET step. In the quenching reaction, the energy differences of ΔΔH ^≠(MCD–MCN) and ΔΔS ^≠(MCD–MCN) at 25 °C were obtained as −1.1 and 0 kJ mol⁻¹, respectively. On the other hand, ΔΔH ^≠(MCD–MCN)=11±2 kJ mol⁻¹ and TΔΔS ^≠(MCD–MCN)=−10±2 kJ mol⁻¹ were given in the thermal back ET reaction. The highest stereoselectivity of 1.7 for [MCD]^·+ found at low temperature (10 °C) was due to the ΔΔS ^≠ value obtained in the thermal back ET reaction. Electronic Supplementary Material Supplementary material is available for this article at and is accessible for authorized users.     

Ca2+-activated K+ channels in the apical membrane ofNecturus choroid plexus

Summary The properties of Ca²⁺-activated K⁺ channels in the apical membrane of theNecturus choroid plexus were studied using single-channel recording techniques in the cell-attached and excised-patch configurations. Channels with large unitary conductances clustered around 150 and 220 pS were most commonly observed. These channels exhibited a high selectivity for K⁺ over Na⁺ and K⁺ over Cs⁺. They were blocked by high cytoplasmic Na⁺ concentrations (110mm). Channel activity increased with depolarizing membrane potentials, and with increasing cytoplasmic Ca²⁺ concentrations. Increasing Ca²⁺ from 5 to 500nm, increased open probability by an order of magnitude, without changing single-channel conductance. Open probability increased up to 10-fold with a 20-mV depolarization when Ca²⁺ was 500nm. Lowering intracellular pH one unit, decreased open probability by more than two orders of magnitude, but pH did not affect single-channel conductance. Cytoplasmic Ba²⁺ reduced both channel-open probability and conductance. The sites for the action of Ba²⁺ are located at a distance more than halfway through the applied electric field from the inside of the membrane. Values of 0.013 and 117mm were calculated as the apparent Ba²⁺ dissociation constants (K _d (0 mV) for the effects on probability and conductance, respectively. TEA⁺ (tetraethylammonium) reduced single-channel current. Applied to the cytoplasmic side, it acted on a site 20% of the distance through the membrane, with aK _d (0 mV)=5.6mm. A second site, with a higher affinity,K _d (0 mV)=0.23mm, may account for the near total block of chanel conductance by 2mm TEA⁺ applied to the outside of the membrane. It is concluded that the channels inNecturus choroid plexus exhibit many of the properties of maxi Ca²⁺-activated K⁺ channels found in other tissues.     

Self-hydroxylation of taurine/α-ketoglutarate dioxygenase: evidence for more than one oxygen activation mechanism

2-Aminoethanesulfonic acid (taurine)/α-ketoglutarate (αKG) dioxygenase (TauD) is a mononuclear non-heme iron enzyme that catalyzes the hydroxylation of taurine to generate sulfite and aminoacetaldehyde in the presence of O₂, αKG, and Fe(II). Fe(II)TauD complexed with αKG or succinate, the decarboxylated product of αKG, reacts with O₂ in the absence of prime substrate to generate 550- and 720-nm chromophores, respectively, that are interconvertible by the addition or removal of bound bicarbonate and have resonance Raman features characteristic of an Fe(III)–catecholate complex. Mutagenesis studies suggest that both reactions result in the self-hydroxylation of the active-site residue Tyr73, and liquid chromatography nano-spray mass spectrometry/mass spectrometry evidence corroborates this result for the succinate reaction. Furthermore, isotope-labeling resonance Raman studies demonstrate that the oxygen atom incorporated into the tyrosyl residue derives from H₂ ¹⁸O and ¹⁸O₂ for the αKG and succinate reactions, respectively, suggesting distinct mechanistic pathways. Whereas the αKG-dependent hydroxylation likely proceeds via an Fe(IV)=O intermediate that is known to be generated during substrate hydroxylation, we propose Fe(III)–OOH (or Fe(V)=O) as the oxygenating species in the succinate-dependent reaction. These results demonstrate the two oxygenating mechanisms available to enzymes with a 2-His-1-carboxylate triad, depending on whether the electron source donates one or two electrons.     

Observations on the distribution of the Gm- and Inv-groups in Hungary

Summary The authors report the distribution of Gm- and Inv-groups in Hungary. In our total material of 378 males and females of different age, the frequency of the following Gm-phenotypes turned out: Gm (1, 2, 4, 5, 10)=4.1%, Gm(1,-2, 4, 5, 10)=25.3%, Gm(1,-2,-4,-5,-10)=3.4%, Gm(1, 2,-4,-5,-10)=0.4%, and Gm(-1,-2, 4, 5, 10)=66.8%. The alleles frequencies have been calculated to be Gm¹=0.1622, Gm^{1, 2}=0.0228, Gm^{4, 5, 10}=0.8150. Observed and expected Gm-phenotypes frequencies are in good agreement under the assumption of the 3-alleles-model Gm¹, Gm^{1, 2}, Gm^{4, 5, 10}. The Invphenotypes frequencies came out to be: Inv(1,-2)=0.5$, Inv(1,2)=8.2%, and Inv(-1,-2)=91.3%.Among our Hungarian samples no significant differences were found, neither in the distribution of Gm-nor Inv-phenotypes and alleles. There exist no significant differences in the distribution of Inv-groups between our Hungarian sample and two German samples, but the Hungarian sample differs clearly from Czechoslovakian and Yugoslavian ones in the distribution of the Gm-phenotypes.Supported by a grant from the Caisse Nationale de Sécurité Sociale.Attaché de Recherche á l'I.N.S.E.R.M.     

Rhodium aryloxide complexes containing terdentate nitrogen ligands, C-O bond formation, hydrogen bonding with phenol and oxidative addition reactions. Molecular structure of an Rh(III)-acetyl complex

The new rhodium(I) phenoxide complexes [Rh(OPh) (2,6-(CH=R²)₂C₅H₃N)] (R² = i-Pr(3), t-Bu(4)) containing strongly electrondonating N-N′-N ligands, have been prepared by a metathesis reaction of [RhCl(2,6-(CH=R²)₂C₅H₃N)] (R² = i-Pr (1), t-Bu (2)) with NaOPh. These rhodium(I) phenoxide complexes 3 and 4, which are very sensitive to O₂ but stable towards H₂O, give with phenol the adducts [Rh(OPh) (2,6-(CH=NR²)₂C₅H₃N)] · HOPh (R2 = i-Pr (5), t-Bu (6)), which contain strong O-HO hydrogen bonds. The hydrogen bonded phenol could not be extracted with diethyl ether, while no exchange of the hydrogen bonded phenol and the phenoxide ligand in 4 is observed on the NMR time scale. However, a small excess of phenol results in exchange of the hydrogen bonded phenol, the coordinated phenoxide ligand and free phenol on the NMR time scale. Reaction of 3 and 4 with p-nitrophenol afforded [Rh(OC₆H₄-(NO₂-4))(2,6-(CH=R²)₂C₅H₃N)] · HOPh (R² = i-Pr (7), t-Bu (8)) in which the formed phenol is hydrogen bonded to the Rh(I)-OC₆H₄-(NO₂-4) moiety. The O-HO bond is less strong than in 5 and 6, as the hydrogen bonded phenol could be removed by diethyl ether.Treatment of 3 with acetyl chloride and benzoyl chloride in benzene at room temperature gave phenylacetate and RhCl₂(C(O)C₆H₃) (2,6(C(H)=N-i-Pr)₂C₅H₃N)] (15), and phenylbenzoate and [RhCl₂(C(O)Ph) (2,6-(C(H)=N-i-Pr)₂C₅H₃N)] (19), respectively. Complex 15 and the analogous complex [RhCl₂(C(O)CH₃) (2,6-(C(H)=N-t-Bu)₂C₅H₃N)] (16) could also be prepared directly from acetyl chloride and 1 or 2, respectively. The single crystal X-ray determination of complex 16, monoclinic, space group P2₁/_c, a = 10.0477(5), b= 11.7268(6), c= 19.2336(9) Å, β = 92.041(4)°, Z = 4, R₁ = 0.0281, shows that the acetyl group occupies an axial position, while the N-N′-N ligand is positioned equatorially. In solution this geometry remains unchanged as was shown by variable temperature ¹H NMR measurements. When the oxidative addition of acetyl chloride to 3 was carried out at −78°C in toluene the intermediate complex [RhCl(OPh) (C(O)Me) (2,6-(C(H)=N-i-Pr)₂C₅H₃N)] (11) could be isolated, which at room temperature reductively eliminates phenylacetate with formation of 1. Oxidative addition of acetyl chlori de to 4 at room temperature gives [RhCl(OPh) (C(O)Me) (2,6-(C(H)=Nt-Bu)₂C₅H₃N)] (12) which yields phenylacetate and 2 at 70°C in benzene by inductive elimination. Treatment of 3 with two equivalents of benzyl chloride afforded a mixture of [RhCl(OPh) (CH₂Ph) (2,6-(C(H)=N-i-Pr)₂C₅H₃N)] (13) and [RhCl₂(CH₂Ph) (2,6-(C(H)=N-i-Pr)₂C₅H₃N)] (17) and some non-characterizable organic products, while 4 only yielded [RhCl(OPh) (CH₂Ph) (2,6-(C(H)=N-tBu)₂C₅H₃N)] (14).     

Genetic,ontogenetic, and tissue-specific variation of dipeptidases in Drosophila melanogaster

Three dipeptidases in Drosophila melanogaster are under independent genetic control and their structural genes have been localized, Dip-A to 2R and Dip-B and Dip-C to 3R (Voelker and Langley, 1978; Ohnishi and Voelker, 1981). These enzymes were characterized with respect to their substrate specificities, genetic variability (electrophoretic mobility and quantitative activity level), ontogeny (activity and isozyme pattern), and tissue localization. The dipeptide substrate specificities of DIP-A and DIP-B overlap each other considerably, but do not overlap with DIP-C. In natural populations, DIP-B and DIP-C are essentially monomorphic electrophoretically whereas DIP-A is polymorphic for three allozymes. Both DIP-A and DIP-B show quantitative genetic variation of activity level within an allozyme class. All three enzymes are expressed at all stages in the life cycle, but DIP-A and DIP-B activities vary considerably according to developmental stage and sex of adult. The tissue localizations of DIP-A and DIP-B activities show similar patterns and a nearly ubiquitous occurrence of both enzymes, but with particularly high values in larval and adult midguts and in the adult female reproductive system. These results suggest a general metabolic role for the enzymes, such as regulation of the concentrated pools of amino acids and oligopeptides found in Drosophila tissues.This work was supported by Public Health Service Grant GM 11546.Paper No. 7066 of the Journal Series of the North Carolina Agricultural Research Service, Raleigh.     

L-alanine:2-oxoglutarate aminotransferase isoenzymes from Arabidopsis thaliana leaves

The occurrence of four l-alanine:2-oxoglutarate aminotransferase (AOAT) isoenzymes (AOAT-like proteins): alanine aminotransferase 1 and 2 (AlaAT1 and AlaAT2, EC 2.6.1.2) and l-glutamate:glyoxylate aminotransferase 1 and 2 (GGAT1 and GGAT2, EC 2.6.1.4) was demonstrated in Arabidopsis thaliana leaves. These enzymes differed in their substrate specificity, susceptibility to pyridoxal phosphate inhibitors and behaviour during molecular sieving on Zorbax SE-250 column. A difference was observed in the electrostatic charge values at pH 9.1 between GGAT1 and GGAT2 as well as between AlaAT1 and AlaAT2, despite high levels of amino acid sequence identity (93 % and 85 %, respectively). The unprecedented evidence for the monomeric structure of both AlaAT1 and AlaAT2 is presented. The molecular mass of each enzyme estimated by molecular sieving on Sephadex G-150 and Zorbax SE-250 columns and SDS/PAGE was approximately 60 kDa. The kinetic parameters: K_m (Ala)=1.53 mM, K_m (2-oxoglutarate)=0.18 mM, k_cat=124.6 s⁻¹, k_cat/K_m=8.1 × 10⁴ M⁻¹·s⁻¹ of AlaAT1 were comparable to those determined for other AlaATs isolated from different sources. The two studied GGATs also consisted of a single subunit with molecular mass of 47.3–70 kDa. The estimated K_m values for l-glutamate (1.2 mM) and glyoxylate (0.42 mM) in the transamination catalyzed by putative GGAT1 contributed to indentification of the enzyme. Based on these results we concluded that each of four AOAT genes in Arabidopsis thaliana leaves expresses different AOAT isoenzyme, functioning in a native state as a monomer.     

Bayesian Estimation in a Generalized Negative Binomial Distribution

A generalized negative binomial (GNB) distribution was introduced by JAIN and CONSUL (1971) and was modified by NELSON (1975). The probability function of the distribution is defined by the function p(x; m, β, θ)= θ^x (1 - θ)^m+βx—x for x=0, 1, …, and zero otherwise, where m>0, 0<θ<1 and β=0 or 1≦β<θ^?1. The Bayes estimators for a number of parametric functions of θ when m and β are known are derived. The prior information on θ may be given by a beta distribution, B(a, b), to which no subjective significance is attached. It has been illustrated that the parameters in the prior distribution can be assigned by a computer. Comparisons are made of the Bayes estimate of P(X=k) to the corresponding ML estimate and the MVU estimate for any given sample to the order n^?1 for different values of k..     

Riboflavin-binding sites associated with flagella of Euglena: A candidate for blue-light photoreceptor?

The hypothesis was tested that reversible riboflavin (RF)-binding sites are part of the photoreceptor in Euglena gracilis. Published evidence shows that the phototactic stimulus — with a flavin-type action spectrum — is perceived at the paraflagellar body (PFB). Flagella with PFBs were isolated from Euglena gracilis by a combined cold and Ca²⁺ shock. Saturable binding of [¹⁴C]RF was demonstrated with such preparations, in the oxidized state as well as under reducing conditions in the presence of dithionite. Affinities for RF were high: K _D (oxidized)=0.08 M, and K _D (reduced)=0.7 M. Flavin mononucleotide and flavin adenine dinucleotide showed lower binding affinities. The in vitro RF binding per unit of protein was enriched approximately tenfold in the flagellar preparations when compared with homogenates of whole cells. The number of (reduced) binding sites per entire flagellum was determined to be in the order of 10⁶. This number is in line with published estimates of chromophores bound in or at the PFB.Abbreviations FAD flavin adenine dinucleotide - FMN flavin mononucleotide - PFB paraflagellar body - RF riboflavin This work was supported by the Deutsche Forscungsgemeinschaft.     

Polymorphisms in the interleukin-4 receptor α chain gene influence susceptibility to HIV-1 infection and its progression to AIDS

Interleukin (IL) 4 is a key T helper-2 cytokine that downregulates and upregulates CCR5 and CXCR4, respectively, the main coreceptors for HIV. Our objective is to investigate whether single-nucleotide polymorphisms (SNPs) in the IL-4 receptor α chain gene (IL4RA) affect HIV infection and its progression to AIDS. The I50V SNP in exon 5 and the haplotypes of six SNPs in exon 12 (E375A, C406R, S411L, S478P, Q551R, and V554I) were studied by polymerase chain reaction and sequencing in 30 HIV⁺ long-term nonprogressors (LTNP), 36 HIV⁺ typical progressors (TP), 55 highly exposed but uninfected individuals (EU), 25 EU-sexuals (EU-Sex; mostly women) and 30 EU-hemophiliacs (EU-Hem; hepatitis C virus⁺), and 97 healthy controls (HC), all Caucasians and lacking CCR5Δ32 homozygosity. V50 homozygosity was increased in LTNP (44%) compared with the other groups [p=0.005; relative risk ratio=3.4, 95% confidence interval (CI)=1.12–10.6, p=0.03]. The most common (C) exon 12 haplotype, ECSSQV, predominated in all groups, but uncommon (U) haplotypes were increased in HIV⁺ individuals (n=64), especially in those (51 of 64) infected via parenteral exposure (35.3%) compared with HC (20.4%) and EU-Hem (18.4%) [p=0.01; odds ratio (OR)=2.14, 95% CI=1.25–3.67, p=0.01]. EU-Sex also had an increased frequency of U-haplotypes (34.8%) (OR=2.10, 95% CI=1.03–4.21, p=0.01) as well as an increased frequency of CU + UU genotypes (60.9%) compared with HC (38.2%) and EU-Hem (26.6%) (p=0.043). Distributions of genotypes fitted Hardy–Weinberg equilibrium. Data suggest that V50 homozygosity associates with slow progression and that exon 12 U-haplotypes might be associated with both susceptibility to infection via parenteral route and resistance to infection via sexual exposure. Further studies are required to confirm these findings.     

Ovarian development rates in the Old World screw-worm fly, Chrysomya bezziana

Rates of ovarian development in relation to temperature were determined for autogenous females of the screw-worm fly, Chrysomya bezziana. Percentage durations of the different ovarian stages (scaled 2–10) were estimated on the basis of observed lengths of the developing oocytes. Mean durations (h) of each ovarian stage was determined at 20, 25, 28 and 35°C. A model of ovarian development rate (%/d) in relation to temperature (T) is presented, the fitted curve being give by R(T)=EXP (–2.73+0.362T–0.0055T²).     

Time course of appearance ofα-bungarotoxin binding sites during development of chick ciliary ganglion and iris

The binding of [¹²⁵I]alpha-bungarotoxin (ABTX) to homogenates of ciliary ganglia and irises from embryonic and posthatching chickens has been examined. Specific, high-affinity binding was found in both tissues [K _D (iris)=2.5 nM;K _D (ganglion)=2.7 nM]. Binding is saturated above 10 nM toxin concentration and is inhibited by low concentrations of the nicotinic antagonistd-tubocurarine. The binding may be associated with a nicotinic cholinergic receptor in both tissues. The amount of binding in the iris begins to increase soon after functional innervation is first observed, at 12 days of incubation (d.i.), and continues to increase up to four months after hatching (a.h.), the oldest age tested. In contrast, ABTX binding in the ciliary ganglion increases fourfold between 7 and 11 d.i., after which the amount of binding remains unchanged up to four months a.h. When compared to the development of choline acetyltransferase (ChAc) and acetylcholinesterase (AChE) activities in the ganglion and iris, ABTX binding follows a pattern similar to that of AChE activity. The largest increases in ChAc activity occur later than those of the postsynaptic markers. After 16 d.i. there are approximately 3×10⁶ toxin molecules bound per neuron in the ciliary ganglion.Submitted by V. A. Chiappinelli in partial fulfillment of the requirements for the PhD degree in the Department of Biobehavioral Sciences, University of Connecticut, Storrs, Connecticut.     

Structure and coordination of CuB in the Acidianus ambivalens aa 3 quinol oxidase heme–copper center

The coordination environment of the Cu_B center of the quinol oxidase from Acidianus ambivalens, a type B heme–copper oxygen reductase, was investigated by Fourier transform (FT) IR and extended X-ray absorption fine structure (EXAFS) spectroscopy. The comparative structural chemistry of dinuclear Fe–Cu sites of the different types of oxygen reductases is of great interest. Fully reduced A. ambivalens quinol oxidase binds CO at the heme a ₃ center, with ν(CO)=1,973 cm⁻¹. On photolysis, the CO migrated to the Cu_B center, forming a Cu_B^I–CO complex with ν(CO)=2,047 cm⁻¹. Raising the temperature of the samples to 25°C did not result in a total loss of signal in the FTIR difference spectrum although the intensity of these signals was reduced sevenfold. This observation is consistent with a large energy barrier against the geminate rebinding of CO to the heme iron from Cu_B, a restricted limited access at the active-site pocket for a second binding, and a kinetically stable Cu_B–CO complex in A. ambivalens aa ₃. The Cu_B center was probed in a number of different states using EXAFS spectroscopy. The oxidized state was best simulated by three histidines and a solvent O scatterer. On reduction, the site became three-coordinate, but in contrast to the bo ₃ enzyme, there was no evidence for heterogeneity of binding of the coordinated histidines. The Cu_B centers in both the oxidized and the reduced enzymes also appeared to contain substoichiometric amounts (0.2 mol equiv) of nonlabile chloride ion. EXAFS data of the reduced carbonylated enzyme showed no difference between dark and photolyzed forms. The spectra could be well fit by 2.5 imidazoles, 0.5 Cl⁻ and 0.5 CO ligands. This arrangement of scatterers would be consistent with about half the sites remaining as unligated Cu(his)₃ and half being converted to Cu(his)₂Cl⁻CO, a 50/50 ratio of Cu(his)₂Cl⁻ and Cu(his)₃CO, or some combination of these formulations. Electronic Supplementary Material Supplementary material is available for this article at .     

Synthesis and crystal structure of the MgCl2(CH3COOC2H5)2· (CH3COOC2H5) adduct

The reaction of α-MgCl₂ with boiling ethyl acetate affords MgCI₂(CH₃COOC₂H₅)₂· (CH₃COOC₂H₅), which is obtained as crystals suitable for X-ray analysis only from the mother liquor. M=315.5, orthorhombic, space group P2₁22₁ (No. 18), a=25.077(3), b=8.616(1), c=7.345(1) Å, V=1587.0(3) ų, Z=4, D_x=1.32 g cm⁻³,λ A(Mo Kα)=0.71069 Å, μ=4.17 cm⁻¹, F(000)=664, T=298 K, observed reflections: 1667, R=0.059 and R_w=0.069. The structure is composed of polymeric chains of MgCl₂(CH₃COOC₂H₅₎₂ and the ethyl acetate molecules occupy a mutually trans position.