Native Molecular Forms of Head Acetylcholinesterase from Adult Drosophila melanogaster: Quaternary Structure and Hydrophobic Character

The native molecular forms of acetylcholinesterase (AChE) present in adult Drosophila heads were characterized by sedimentation analysis in sucrose gradients and by nondenaturing electrophoresis. The hydrophobic properties of AChE forms were studied by comparing their migration in the presence of Triton X100, 10-oleyl ether, or sodium deoxycholate, or in the absence of detergent. We examined the polymeric structure of AChE forms by disulfide bridge reduction. We found that the major native molecular form is an amphiphilic dimer which is converted into hydrophilic dimer and monomer on autolysis of the extracts, or into a catalytically active amphiphilic monomer by partial reduction. The latter component exists only as trace amounts in the native enzyme. Two additional minor native forms were identified as hydrophilic dimer and monomer. Although a significant proportion of AChE was only solubilized in high salt, following extractions in low salt, this high salt-soluble fraction contained the same molecular forms as the low salt-soluble fractions: thus, we did not detect any molecular form resembling the asymmetric forms of vertebrate cholinesterases.     

Molecular Properties of Drosophila Acetylcholinesterase

Abstract: Two distinct classes of acetylcholinesterase (AChE) from the fruit fly Drosophila melanogaster are reported: a soluble species that shows heterogeneity of forms and a particulate species. The subunit composition of the particulate enzyme was studied using the active site label [³H]diisopropylfluorophosphate. Comparison of the electrophoretic patterns on nondenaturing gels using the activity stain and the active site label shows that the label is specific to AChE. The smallest active site-containing subunit of the enzyme is a monomer of $60,000 daltons MW. Two such units are linked by disulphide bonds to produce a dimer of about 110,000 daltons. Another monomeric form of MW $64,000 daltons, although present, does not participate in the dimerisation. The particulate enzyme when solubilised exists as a 9–10S species as determined by sucrose gradient centrifugation. This species has a MW>200,000, as shown by its behaviour on a coarse-bead Sephadex-G200 column. Electrophoretic analysis suggests a MW of nearly 250,000 daltons for this form. Thus, this species is likely to be a tetramer. One possibility is that this tetramer is made up of two units of 64,000 daltons each and a dimer of 110,000 daltons. Preliminary data on mutant enzymes that support such a possibility are also presented.     

Comparison of a Homology Built Model of Angiogenin to its Crystal Structure

The comparison of our homology built model of human angiogenin with the recently determined x-ray structure of the same is reported. The basic details of the structure in terms of alpha -helices and beta sheets were found to be common. The main differences between the model and the x-ray data lie in a C-terminal rearrangement in the x-ray structure that causes the C-terminus to end in a 3₁₀ helix which puts the residue GLN-117 (ALA-122 in bovine pancreatic ribonuclease A, RNaseA) into the active site. The homology model was updated by producing a new sequence alignment using the information from the x-ray data which improved the r.m.s. by 0.5Å. This new alignment is also reported here. A check for systematic bias was carried out using the RNaseA structures from which the x-ray and homology models were derived. A detailed comparison of torsion angles and hydrogen bonding between all the structures have been compared and the model displays several hydrogen bonds that are not present in the parent RNaseA structures but are present in the x-ray structure of angiogenin.Electronic Supplementary Material available.     

环境对果蝇P转座因子的作用关系研究

转座因子,重组、整合、遗传效应等是目前遗传学领域的一个研究热题。转座因子对遗传变异、宗系进化、突变频率、物种形成、新基因的产生以及对分子生物学、遗传工程学、群体遗传学和数量遗传学等方面的研究都有着重要的意义,主要对果蝇的P转座因子以及环境对P转座因子遗传效应的作用关系进行了研究。     

Structural Evaluation of Distant Homology. A 3-D Model of the Ligand Binding Domain of the Nicotinic Acetylcholine Receptor Based on Acetylcholinesterase: Consistency with Experimental Data

Acetylcholine is a ligand for both acetylcholinesterases and nicotinic acetylcholine receptors. Hence, at least some local sequence and structural similarities between the acetylcholinesterases and the receptors which recognize acetylcholine (ACh) might be expected. Peterson [2] produced an alignment of the ACh binding region between these two types of ACh–binding molecules, featuring a number of well conserved residues. The extent of this region of sequence similarity suggests the possible existence of a common ancenstral ACh binding module. To attempt to further validate Petersons sequence alignment we have built a homology model of the ACh binding domain of the human neuromuscular nicotinic acetylcholine receptor based on the structure of acetylcholinesterase from Torpedo californica. Using this 3–D model we have examined the residues which were previously shown to interact with the endogenous ligand by various methods (mapping, site–directed mutagenesis). The consistency of such data with the model provides further support for a structural similarity and possibly a divergent evolutionary relationship between the ACh–binding domains of these two classes of proteins. Results suggest that this model may be able to contribute to an understanding of the structure and function of the ACh receptor. Using this case as an example, we propose that 3–dimensional computer modeling can be used as a tool to evaluate distant homologies when adequate experimental data (e.g., site–directed mutagenesis) is available.Supplementary material to this paper is available in electronic form at http://dx.doi.org/10.1007/s0089460020046     

果蝇基因组中内含子数目随其长度的分布研究

为了研究内含子可能储存的有关生命进化信息,本研究以果蝇基因组中的内含子为样本,统计了内含子在不同长度区间内的相对频数,得到了内含子随其长度的分布,发现这种分布呈现出一定的规律性,即长度在1~80 bp内的内含子数目随其长度增加而增加,长度大于80 bp的内含子数目随其长度的增加而减少;本研究推论这种分布规律应该与某种分布模型一致,经过各种分布的拟合,最后发现这种分布与Γ(α,β)分布相符合;另外,将果蝇内含子分布规律和相应的外显分布进行了比较分析,发现虽然两类序列的分布规律都符合Γ(α,β)分布,但也存在明显的区别,并就此讨论了两类序列在生物进化中的意义。     

DMAP-85: A τ-Like Protein from Drosophila melanogaster Larvae

Abstract: Microtubule-associated proteins (MAPs) play major regulatory roles in the organization and integrity of the cytoskeletal network. Our main interest in this study was the identification and the analysis of structural and functional aspects of Drosophila melanogaster MAPs. A novel MAP with a relative molecular mass of 85 kDa from Drosophila larvae was found associated with taxol-polymerized microtubules. In addition, this protein bound to mammalian tubulin in an overlay assay and coassembled with purified bovine brain tubulin in microtubule sedimentation experiments. The estimated stoichiometry of 85-kDa protein versus tubulin in the polymers was 1:5.3 ± 0.2 mol/mol. It was shown that the 85-kDa protein bound specifically to an affinity column of Sepharose-βII-(422–434) tubulin peptide, which contains the sequence of the MAP binding domain on βII-tubulin. Affinity-purified 85-kDa protein enhanced microtubule assembly in a concentration-dependent manner. This effect was significantly decreased by the presence of the βII-(422–434) peptide in the assembly assays, thus confirming the specificity of the 85-kDa protein interaction with the C-terminal domain on tubulin. Furthermore, this protein also exhibited a strong affinity for calmodulin, based on affinity chromatographic assays. Monoclonal and polyclonal anti-τ antibodies, including sequence-specific probes that recognize repeated microtubule-binding motifs on τ, MAP-2, and MAP-4 and specific N-terminal sequences of τ, cross-reacted with the 85-kDa protein from Drosophila larvae. These results suggest that τ and Drosophila 85-kDa protein share common functional and structural epitopes. We have named this protein as DMAP-85 for Drosophila MAP. The finding on a Drosophila protein with functional homology and structural similarities to mammalian τ opens new perspectives to understand the cellular roles of MAPs.     

Prolongation of Life in an Experimental Model of Aging in Drosophila Melanogaster

(R)-Deprenyl, the archetypical monoamine oxidase-B inhibitor, has been shown to increase life-span in a number of species. Although many theories for this effect have been suggested, for example, an increase in superoxide dismutase (SOD) activity, the mechanism of action has yet to be elucidated. To investigate this phenomenon, we have examined the effects of (R)-deprenyl, and some aliphatic propargylamines, in an experimental aging model in Drosophila melanogaster. Both wild-type Oregon-R type flies, as well as a SOD knock-out mutant strain were used. Flies obtained from a series of paired mates were divided equally among treatment groups. In all studies, flies were treated for the duration of life following adult emergence. The aging model consists of substitution of sucrose with galactose in the regular food media of the flies. Initial experiments confirmed that such a substitution resulted in a significant (p < 0.01, Breslow test) reduction in mean and maximal life-span of flies, an effect not due to nutrient deprivation. Inclusion of (R)-deprenyl and the aliphatic propargylamines in the media, at average daily doses in the range 0.5-1ng/fly/day, led to a significant increase in mean and maximal life-span of galactose-treated, but not control flies. This effect was seen in both wild-type and mutant flies.     

Abdominal pigmentation in Drosophila melanogaster females from natural Indian populations

Females of Drosophila melanogaster collected from five geographically distant populations in India were analysed for the intensity of pigmentation in the 5th, 6th and 7th segments of the abdomen. In all three segments, this intensity was found to vary among individuals of any given population, and, furthermore, different populations differ with respect to this phenotypic trait. Statistical analysis revealed significant intra- and interpopulational variation. A clinical pattern was detected: females from populations closer to the equator tended to have lighter cuticle, in which case differences between the three segments could not be detected and all three segments responded both independently and jointly to latitudinal variation, as indicated by a statistically significant positive correlation between latitude and pigmentation score. This is the first report on abdominal pigmentation analysis in natural populations of D. melanogaster that provides evidence that phenotypic flexibility reflects temperature differences, as a result of which abdominal pigmentation shows geographic differentiation.     

Population Dynamics of Herpetomonas ampelophilae, with a Note on the Systematics of Herpetomonas from Drosophila spp.

SYNOPSIS Previous surveys of feral Drosophila melanogaster in Clarke County, Georgia, indicated 10 to 30% of the flies were parasitized with Herpetomonas ampelophilae. In the present study, herpetomoniasis in D. melanogaster reared in population cages rose from 0.01% to 90% in 14 days. The infection fluctuated between 5% and 100% during the 250 days in which the ageledeme was maintained.
Herpetomonas were found endotrophically, peritrophically, and in the Malpighian tubules. In population cages, although there were sporadic rises in percentages, peritrophic and Malpighian tube infections usually remained low. Endotrophic infections often reached 100%, with individual flies being heavily parasitized. Although Chatton felt that several species of Herpetomonas inhabit a single species of Drosophila , we suggest that all the Herpetomonas in D. melanogaster be referred to as H. ampelophilae.     

Simulation of gene pyramiding in Drosophila melanogaster

Gene pyramiding has been successfully practiced in plant breeding for developing new breeds or lines in which favorable genes from several different lines were integrated.But it has not been used in animal breeding,and some theoretical investigation and simulation analysis with respect to its strategies,feasibility and efficiency are needed before it can be implemented in animals.In this study,we used four different pure fines of Drosophila melanogaster,each of which is homozygous at a specific mutant gene with a visible effect on phenotype,to simulate the gene pyramiding process and analyze the duration and population size required in different pyramiding strategies.We finally got the ideal individuals,which are homozygous at the four target genes simultaneously.This study demonstrates that gene pyramiding is feasible in animal breeding and the interaction between genes may affect the final results.     

紫外线对黑尾果蝇的生物学效应

野生型黑尾果蝇经紫外线连续处理3代,随着照射时间延长,每代均表现显著的生物学效应。这些生物学效应主要是子代数量减少,羽化延续时间缩短,体重减轻,而且有致略效应和间接的诱变效应。但处理后的子代性别比例变化不大。     

Simulation of gene pyramiding in Drosophila melanogaster

Gene pyramiding has been successfully practiced in plant breeding for developing new breeds or lines in which favorable genes from several different lines were integrated.But it has not been used in animal breeding,and some theoretical investigation and simulation analysis with respect to its strategies,feasibility and efficiency are needed before it can be implemented in animals.In this study,we used four different pure lines of Drosophila melanogaster,each of which is homozygous at a specific mutant gene ...     

Intron length evolution in Drosophila

I present data on the evolution of intron lengths among 3 closely related Drosophila species, D. melanogaster, Drosophila simulans, and Drosophila yakuba. Using D. yakuba as an outgroup, I mapped insertion and deletion mutations in 148 introns (spanning approximately 30 kb) to the D. melanogaster and D. simulans lineages. Intron length evolution in the 2 sister species has been different: in D. melanogaster, X-linked introns have increased slightly in size, whereas autosomal ones have decreased slightly in size; in D. simulans, both X-linked and autosomal introns have decreased in size. To understand the possible evolutionary causes of these lineage- and chromosome-specific patterns of intron evolution, I studied insertion-deletion (indel) polymorphism and divergence in D. melanogaster. Small insertion mutations segregate at elevated frequencies and enjoy elevated probabilities of fixation, particularly on the X chromosome. In contrast, there is no detectable X chromosome effect on fixations in D. simulans. These findings suggest X chromosome-specific selection or biased gene conversion-gap repair favoring insertions in D. melanogaster but not in D. simulans. These chromosome- and lineage-specific patterns of indel substitution are not easily explained by existing general population genetic models of intron length evolution. Genomic data from D. melanogaster further suggest that the forces described here affect introns and intergenic regions similarly.     

果蝇蛋白质组学研究进展

蛋白质组学旨在阐明基因组所表达的真正执行生命活动的全部蛋白质的表达规律和生物功能。随着人类基因组学计划的逐渐成熟,分子水平的实验技术不断发展,蛋白质组学的研究被提高到了前所未有的高度。果蝇是生命科学领域最为常用的一种模式生物,长期的系统研究也使果蝇的基因组成为至今注释最好的基因组之一,为功能基因组研究奠定了基础。但由于技术的限制,迄今有关果蝇蛋白质组学研究的报道尚不多见。近年来果蝇蛋白质组学的研究主要包括表达谱、修饰谱、比较蛋白质组学和疾病模型蛋白质组等四个方向,为进一步开展人类疾病临床蛋白质组学研究奠定了基础。     

Cytological location and further characterization of the Third chromosome resistance gene in Drosophila melanogaster

Mutations in the Third chromosome resistance (Tcr; 3-39.6) gene confer dominant resistance to α-methyl dopa and suggest the gene is involved in catecholamine metabolism. Evidence for involvement in catecholamine metabolism comes from the three phenotypes associated with the mutant Tcr chromosomes dominant resistance, dominant rescue of partially complementing l(2)amd alleles, and recessive lethal phenotypes. Only dominant resistance to αs-methyl dopa, however, was mapped to the Tcr locus. Both recessive lethality and dominant rescue of l(2)amd alleles have now been mapped to the Tcr gene and, through the isolation of a new deletion in the region, we demonstrate these phenotypes are due to a loss of Tcr function. This deletion places the Tcr gene in the 69B4-5 to 69C8-11 region. Additionally, we have tested and verified three predictions of the biochemical model proposed by Bishop, Sherald, and Wright (1989) for the function of the Tcr protein. This revised version was published online in July 2006 with corrections to the Cover Date.     

Topological properties of protein-protein and metabolic interaction networks of Drosophila melanogaster

Introduction The concept of networking and its omnipresent na- ture has been an issue of considerable curiosity, cap- tivating scientists worldwide over the past few years. Simple networking can be witnessed in the physical connectivity of computer terminals or routers. On broader perspective, many networks, such as social ties—familial and professional, World Wide Web, net- work of scientific papers connected by citations, elec- trical power grids, transportation systems, and biolog- ical n…     

Inferring the effects of demography and selection on Drosophila melanogaster populations from a chromosome-wide scan of DNA variation

Identifying regions of the Drosophila melanogaster genome that have been recent targets of positive Darwinian selection will provide evidence for adaptations that have helped this species to colonize temperate habitats. We have begun a search for such genomic regions by analyzing multiple loci (about 250) dispersed across the X chromosome in a putatively ancestral population from East Africa and a derived European population. For both populations we found evidence for past changes in population size. We estimated that a major bottleneck associated with the colonization of Europe occurred about 3,500-16,000 years ago. We also found that while this bottleneck can account for most of the reduction in variation observed in the European sample, there is a deficit of polymorphism in some genomic regions that cannot be explained by demography alone.