Hemoglobin as a potential source of natural regulatory oligopeptides

Theoretical structure-function analysis of all possible hemoglobin molecule fragments was performed to determine sites that could be potential sources of regulatory oligopeptides. Known data on bovine hemoglobin primary structure and information of the EROP-Moscow database concerning structure and functions of natural oligopeptides were used along with a computer program complex. A total of 6750 natural non-hemoglobin oligopeptides with hemoglobin fragments of 2–14 amino acid residues were found. Structures of 20 of them were completely identical to hemoglobin fragments. Most of the revealed oligopeptides exhibit properties of neuropeptides, antimicrobial agents, and hormones. A number of them exhibit functions previously not known for hemoglobin fragments. The possibility of natural formation of regulatory oligopeptides from hemoglobin and other food protein molecules, generation of the exogenous oligopeptide pool, their participation in regulation processes as well as accordance of results obtained here with the oligopeptide continuum concepts are discussed.     

Biochemical problems of regulation by oligopeptides

Some problems are considered which arise in biochemical studies on structure and function of natural oligopeptides consisting of 2–50 amino acid residues. The problems under consideration include the generation of oligopeptides from precursors, chemical structure, the role of functionally important radicals and spatial configuration, and structure-function relationships. Different types of regulation are shown mainly for oligopeptides involved in muscle contraction.Translated from Biokhimiya, Vol. 69, No. 11, 2004, pp. 1565–1573.Original Russian Text Copyright © 2004 by Zamyatnin.     

Allatotropin, leucokinin and AKH in honey bees and other Hymenoptera

In the honey bee no allatotropin gene has been found, even though allatotropin stimulates the synthesis of juvenile hormone in this species. We report here that honey bees and other Hymenoptera do have a typical allatotropin gene, although the peptides predicted have a somewhat different structure from that of other insect allatotropins. Polyclonal antisera to honey bee allatotropin reacted with material in the neurohemal organs of the segmental nerves of abdominal ganglia. We were unable to find the allatotropin peptide using mass spectrometry in extracts from these tissues. Thus the expression of this gene in honey bees is less important than in other insect species. We also characterized the leucokinin gene which similarly appears to be very weakly expressed in worker honey bees. Unlike the allatotropin gene, which is conserved within Hymenoptera, the leucokinin gene is much more variable in structure and was not found in ants nor the parasitic wasp Nasonia vitripennis. The absence of significant expression of adipokinetic hormone (AKH) in the honey bee may be due to the existence of a second TATA box in the promotor region of the gene, which explains the production of an mRNA encoding a putative peptide precursor from which no AKH should be released. Such a second TATA box was not found in other Hymenoptera, and may therefore be specific for the two Apis species. It is suggested that functional disintegration of this important metabolic gene became possible in Apis because of the highly evolved social nature of the species.     

Preliminary Screening of Microorganisms Producing Succinic Acid from n-Paraffin

Screening tests in search for microorganisms capable of producing succinic acid from n-paraffin were carried out. Most of the microorganisms that accumulated succinic acid in culture broth when incubated in the media containing super heavy n-paraffin as the carbon source were found to belong to the genus Candida. The largest quantity of succinic acid production from n-paraffin, 4160 μ/ml, was obtained with a strain, Candida brumptii IFO 0731.     

Partial characterization and isolation of earwig `allatostatins'': biological activities in earwigs and cockroaches

Allatostatins are a family of neuropeptides first isolated from the cockroach, Diploptera punctata, that inhibit juvenile hormone production in that species (but do not do so in earwigs), and inhibit hindgut muscle contractions in some insects, including the earwig, Euborellia annulipes. We examined whether material from earwig brains is similar to cockroach allatostatins biochemically, immunologically and physiologically. Brain extracts from adult female earwigs were separated by high performance liquid chromatography (HPLC), followed by radioimmunoassay using antibodies to cockroach allatostatin (Dip-AST). Fractions that co-eluted with cockroach allatostatins were immunoreactive, and at least two peaks of immunoreactivity were detected. Material from each peak at 10 nM Dip-AST equivalents inhibited juvenile hormone biosynthesis in vitro by corpora allata of 2-day virgin D. punctata cockroaches; 1 nM was less effective, and non-immunoreactive fractions failed to inhibit juvenile hormone biosynthesis. Both crude and Sep-Pak (Waters) purified extracts of brains of earwigs containing 1 nM Dip-AST equivalents failed to suppress hindgut contractions in vitro of 2-day earwigs and of brooding female earwigs. In contrast, 1 nM cockroach allostatin 1 (Dip-AST 7) reversibly inhibited hindgut contractions in vitro. These results suggested the presence of another brain factor, such as proctolin, that counteracts the inhibitory effects of Dip-AST. In support of this hypothesis, proctolin stimulated hindgut contractions in vitro at 1 nM; the effects of equal concentrations of allatostatin and proctolin varied with the stage of the female. Furthermore, HPLC-separated fractions that co-eluted with cockroach allatostatin and were immunoreactive with antibodies to Dip-AST suppressed hindgut contractions in vitro of 2-day female earwigs. Finally, crude brain extracts of earwigs suppressed earwig juvenile hormone biosynthesis in vitro in glands of low, but not in glands of high, activity. Thus, earwig brain extract after HPLC separation has Dip-AST-like material that inhibits cockroach corpora allata and suppresses earwig hindgut contractions. Sep-Pak-extracted earwig brain material, however, does not inhibit earwig gut contraction. Although synthetic Dip-AST 7 does not inhibit juvenile hormone synthesis by earwig corpora allata, there is heat-stable material in earwig brain extract that does have this action.     

Software for analysis of bacterial genomes

GenomeExplorer is a program for comparative analysis of regulation in prokaryotic genomes. The program has options for signal search, comparison of gene samples, search for paralogs and orthologs, iterative construction of signal profiles. The program has a convenient graphic interface, allowing for navigation in the annotation window, in the genome map, and in the table of gene similarities. The use of the system clipboard allows one to export the results of analysis into Word and Excel, and to call external programs via the Internet.     

Database on pharmacophore analysis of active principles, from medicinal plants

Plants continue to be a major source of medicines, as they have been throughout human history. In the present days, drug discovery from plants involves a multidisciplinary approach combining ethnobotanical, phytochemical and biological techniques to provide us new chemical compounds (lead molecules) for the development of drugs against various pharmacological targets, including cancer, diabetes and its secondary complications. In view of this need in current drug discovery from medicinal plants, here we describe another web database containing the information of pharmacophore analysis of active principles possessing antidiabetic, antimicrobial, anticancerous and antioxidant properties from medicinal plants. The database provides the botanical, taxonomic classification, biochemical as well as pharmacological properties of medicinal plants. Data on antidiabetic, antimicrobial, anti oxidative, anti tumor and anti inflammatory compounds, and their physicochemical properties, SMILES Notation, Lipinski's properties are included in our database. One of the proposed features in the database is the predicted ADMET values and the interaction of bioactive compounds to the target protein. The database alphabetically lists the compound name and also provides tabs separating for anti microbial, antitumor, antidiabetic, and antioxidative compounds. AVAILABILITY: http://www.hccbif.info /     

南方鲇两种生长激素受体的结构分析及其组织分布和激素调节

采用生物信息学方法,从东方纯（Fugurubripes）基因组中分离出两种不同基因编码的GHR1（fGHR1）和GHR2（fGHR2）。随后设计引物,采用RT-PCR和Smart^TM Race相结合的方法从南方鲇（Silurus meridionalis）肝脏中克隆出两种不同基因编码的生长激素受体,即GHR1（scGHR1）和GHR2（scGHR2）cD-NA,其全长分别为1985bp（编码602个氨基酸）和2300bp（编码553个氨基酸）。这两种受体都含有生长激素受体典型的标志性基序FGDFS,以及细胞内属于细胞因子受体家族共有的Box1和Box2序列框。同时两者又存在差异,表现在胞外半胱氨酸残基和胞内酪氨酸残基数目的不同。采用半定量RT-PCR方法研究了scGHR1和scGHR2在南方鲇各组织中表达量的差异,结果表明：scGHRl和scGHR2有广泛的组织分布,在肝脏中两者的表达量最高。用17p雌二醇（E2）、17α-甲基睾酮（MT）和可的松（conisol）对南方鲇进行药物处理,结果表明E2能够下调肝脏scGHR1和scGHR2 mRNA水平,而MT上调scGHR1和scGHR2 mRNA水平。同时,经cortisol处理后scGHR1 mRNA表达水平上升,而scGHR2 mRNA表达水平不变。     

An analysis of historical trends in classical biological control of arthropods suggests need for a new centralized database in the USA

The documentation of biological control agents targeting arthropods in the United States has historically been subject to less regulation relative to weed biological control releases. This study reviews publicly available databases to track environmental releases of biological control agents targeting arthropods in the United States. It then presents available data for the states with the most releases between 1962 and 2005: Hawaii, California and Florida. These data indicate a clear decline in rates of introduction since 1982 or 1994, depending on the source. Existing record-keeping systems offer incomplete or inconsistent data for evaluation because they were designed with limited goals, attempt to capture excessive detail and are thus impractical, or are insufficiently resourced. Existing databases cannot be used to answer meaningful questions regarding non-target effects of introduced control agents. Current databases are inappropriately designed and insufficiently resourced to meet today's research and regulatory needs. We propose and describe a new database system for classical biological control of arthropods.     

Methyl-Typing: An improved and visualized COBRA software for epigenomic studies

Combined bisulfite restriction analysis (COBRA) is one of the most commonly used methylation quantification methods. However, it focuses on relatively few restriction enzymes. Here, we present Methyl-Typing, a web-based software that provides restriction enzyme mining data for methyl-cytosine-containing sequences following bisulfite-conversion. Gene names, accession numbers, sequences, PCR primers, and file upload are accessible for input. Promoter sequences and restriction enzymes for CpG- and GpC-containing recognition sites are retrieved. Four representative enzymes were tested successfully by COBRA on the experimental work. Therefore, the Methyl-Typing tool provides a comprehensive COBRA-restriction enzyme mining. It is freely available at http://bio.kuas.edu.tw/methyl-typing.     

State of the NPY-ergic System of the Hypothalamic Arcuate Nucleus in Rats with Experimentally Induced Diabetes Mellitus

We have carried out a quantitative study of the state of a neuropeptide Y (NPY)-ergic system of the hypothalamic arcuate nucleus (AN) in rats in the norm and in streptozotocin (STZ)-induced diabetes mellitus. The NPY-containing objects were identified using an indirect immunofluorescent technique; a system of digital analysis of images and a special software were used. Analysis of the morphometric parameters of the NPY-immunopositive objects within the AN sections allowed us to classify in an automatic mode such objects as neurons of various sizes, fibers, and terminals, as well as to count the number of objects of each class. In addition, the fluorescent intensity of the objects under study (directly proportional to the concentration and amounts of NPY in these objects) was determined. On the basis of these parameters, criteria of the state of synthesis and secretion of NPY and NPY-ergic innervation in the AN subnuclei were proposed. In control animals, the greatest number of NPY-immunopositive neurons and total content of NPY in the neurons and terminals were observed in the ventrolateral and dorsomedial subnuclei of the AN (vl- and dmAN, respectively); this fact points to the high intensities of NPY synthesis and secretion in these structures. The greatest content of NPY in the nerve terminals within the vlAN zone is indicative of the highest intensity of NPY-ergic afferent influences on this subnucleus. It is concluded that the approach we proposed is adequate enough for integral quantitative estimation of the state of peptidergic systems. The development of STZ-induced diabetes mellitus caused changes in the NPY-ergic system in different AN subnuclei; such changes demonstrated certain specificity. In particular, an increase in the number of immunopositive neurons and the total content of NPY in the neurons of vlAN and the ventromedial AN subnucleus (vmAN), which is evidence in favor of intensification of synthesis of the neuropeptide, was not accompanied by an adequate rise in the NPY content in the terminals located in the subnucleus under study. The content of NPY in the eminentia medianus also decreased. The above data allow us to hypothesize that the NPY-ergic system of the AN under conditions of diabetes mellitus is functionally insufficient.     

Relationship between leptin receptor and polycystic ovary syndrome

Polycystic ovary syndrome (PCOS) is one of the most common endocrine disorders, which is involved in the multi-system disease, and its etiology is still not clearly understood. It is currently considered that not only the genetic factors but also the environment factors play a crucial role in the pathogenesis of PCOS. Obesity plays an important role through the insulin, leptin and endocannabinoid system in the pathological process of PCOS, leading to more severe clinical manifestations. The aim of our present study is to investigate whether there is association between single nucleotide polymorphisms (SNPs) of Gln223Arg and Pro1019Pro in the leptin receptor gene (LEPR) and PCOS in a Korean population. Interestingly, a significant association was found between the Pro1019Pro in LEPR gene and PCOS, and a highly significant association was found between the Gln223Arg in LEPR gene and PCOS (P = 0.033, OR = 1.523, 95% confidence interval and P < 0.0001, OR = 0.446, 95% confidence interval). Moreover, genotype combination and haplotype analyses indicate that Gln223Arg and Pro1019Pro polymorphisms of LEPR are significantly associated with the risk of PCOS.     

Single nucleotide polymorphisms in genes that are common targets of luteotropin and luteolysin in primate corpus luteum: Computational exploration

Luteal insufficiency affects fertility and hence study of mechanisms that regulate corpus luteum (CL) function is of prime importance to overcome infertility problems. Exploration of human genome sequence has helped to study the frequency of single nucleotide polymorphisms (SNPs). Clinical benefits of screening SNPs in infertility are being recognized well in recent times. Examining SNPs in genes associated with maintenance and regression of CL may help to understand unexplained luteal insufficiency and related infertility. Publicly available microarray gene expression databases reveal the global gene expression patterns in primate CL during the different functional state. We intend to explore computationally the deleterious SNPs of human genes reported to be common targets of luteolysin and luteotropin in primate CL. Different computational algorithms were used to dissect out the functional significance of SNPs in the luteinizing hormone sensitive genes. The results raise the possibility that screening for SNPs might be integrated to evaluate luteal insufficiency associated with human female infertility for future studies.     

Expression and genomic imprinting of the porcine Rasgrf1 gene

Imprinted genes play important roles in mammalian growth, development and behavior. The Rasgrf1 (Ras protein-specific guanine nucleotide exchange factor 1) gene has been identified as an imprinted gene in mouse and rat. In the present study, we detected its sequence, imprinting status and expression pattern in the domestic pigs. A 228 bp partial sequence located in exon 14 and a 193 bp partial sequence located in exon 1 of the Rasgrf1 gene in domestic pigs were obtained. A G/A transition, was identified in Rasgrf1 exon 14, and then, the reciprocal Berkshire × Wannan black F₁ hybrid model and the RT-PCR-RFLP method were used to detect the imprinting status of porcine Rasgrf1 gene at the developmental stage of 1-day-old. The expression profile results indicated that the porcine Rasgrf1 mRNA was highly expressed in brain, pituitary and pancreas, followed by kidney, stomach, lung, testis, small intestine, ovary, spleen and liver, and at low levels of expression in longissimus dorsi, heart, and backfat. The expression levels of Rasgrf1 gene in brain, pituitary and pancreas tissues were significantly different between the two reciprocal F₁ hybrids. Imprinting analysis showed that porcine Rasgrf1 gene was maternally expressed in the liver, small intestine, paternally expressed in the lung, but biallelically expressed in brain, heart, spleen, kidney, stomach, pancreas, backfat, testis, ovary, longissimus dorsi and pituitary tissues.     

Multiorgan autoimmunity in a Turner syndrome patient with partial monosomy 2q and trisomy 10p

Turner syndrome is a condition caused by numeric and structural abnormalities of the X chromosome, and is characterized by a series of clinical features, the most common being short stature and gonadal dysgenesis. An increased frequency of autoimmune diseases as well as an elevated incidence of autoantibodies has been observed in Turner patients.     

High-throughput sequence analysis of small RNAs in skotomorphogenic seedlings of Brassica rapa ssp. rapa

Skotomorphogenic development is the process by which seedlings adapt to a stressful dark environment. Such metabolic responses to abiotic stresses in plants are known to be regulated in part by microRNAs (miRNAs); however, little is known about the involvement of miRNAs in the regulation of skotomorphogenesis. To identify miRNAs at the genome-wide level in skotomorphogenic seedlings of turnip (Brassica rapa subsp. rapa), an important worldwide root vegetable, we used Solexa sequencing to sequence a small RNA library from seedlings grown in the dark for 4 days. Deep sequencing showed that the small RNAs (sRNAs) were predominantly 21 to 24 nucleotides long. Specifically, 13,319,035 reads produced 359,531 unique sRNAs including rRNA, tRNA, miRNA, small nuclear RNA (snRNA), small nucleolar RNA (snoRNA), and unannotated sRNAs. Sequence analysis identified 96 conserved miRNAs belonging to 36 miRNA families and 576 novel miRNAs. qRT-PCR confirmed that the miRNAs were expressed during skotomorphogenesis similar to the trends shown by the Solexa sequencing results. A total of 2013 potential targets were predicted, and the targets of BrmiR157, BrmiR159 and BrmiR160 were proved to be regulated by miRNA-guided cleavage. These results show that specific regulatory miRNAs are present in skotomorphogenic seedlings of turnip and may play important roles in growth, development, and response to dark environment.     

Metabolite induction via microorganism co-culture: A potential way to enhance chemical diversity for drug discovery

Microorganisms have a long track record as important sources of novel bioactive natural products, particularly in the field of drug discovery. While microbes have been shown to biosynthesize a wide array of molecules, recent advances in genome sequencing have revealed that such organisms have the potential to yield even more structurally diverse secondary metabolites. Thus, many microbial gene clusters may be silent under standard laboratory growth conditions. In the last ten years, several methods have been developed to aid in the activation of these cryptic biosynthetic pathways. In addition to the techniques that demand prior knowledge of the genome sequences of the studied microorganisms, several genome sequence-independent tools have been developed. One of these approaches is microorganism co-culture, involving the cultivation of two or more microorganisms in the same confined environment. Microorganism co-culture is inspired by the natural microbe communities that are omnipresent in nature. Within these communities, microbes interact through signaling or defense molecules. Such compounds, produced dynamically, are of potential interest as new leads for drug discovery. Microorganism co-culture can be achieved in either solid or liquid media and has recently been used increasingly extensively to study natural interactions and discover new bioactive metabolites. Because of the complexity of microbial extracts, advanced analytical methods (e.g., mass spectrometry methods and metabolomics) are key for the successful detection and identification of co-culture-induced metabolites.