Search for evolutionary ancient, relict, regulatory peptides

Among numerous regulatory peptides (RP) it is possible to presumably indicate the relict, evolutionary ancient RP. They combine three features: formation from non-specialized peptides-precursors, a comparatively high resistance to action of proteases in the organism media, and maximal simplicity of their structure. The examples of them are glyprolines—a recently identified RP family, as well as tuftsin. Several other praline-containing RP in terminal sites also seem to belong to the evolutionary ancient RP. The proposed approach to studies on the RP evolution is additional to those used traditionally in this problem.     

Search for ancient patterns in protein sequences

Proteins of related functions are often similar in sequence, reflecting a common phylogenetic origin. Proteins with no known homology are probably diversified proteins, too distantly related to known sequences in databases to retain significant similarity. All proteins, however, probably share common ancestries if one moves far enough back in evolution; therefore, given the huge accumulation of protein sequences in current databases, it could be expected that some proteins with no obvious sequence resemblance to any other share some residues that could represent footprints of ancient common ancestries. To identify such putative footprints, we have searched for short stretches of amino acids present in a given protein sequence that are also found in a significant number of nonrelated proteins in the database. The significantly high frequency of occurrence of these patterns in the database would support a common evolutionary source, and a diversity of non-related proteins that contain the pattern would express their ancient origin. Using this strategy, significant patterns were found in actual exons, but not in randomized amino acid sequences, nor in translated sequences of noncoding DNA, suggesting that this strategy actually leads to the identification of patterns with a biological significance. These significant patterns are not randomly positioned along the sequences analyzed, but they tend to accumulate within specific regions, producing a profile of discrete domains. In some well-known proteins analyzed in this study, some of these domains are coincident with known motifs. Thus, the procedure described in this paper could be useful for identifying ancient patterns and domains in protein sequences, some of which could also have a functional or structural significance.     

Functional continuum of the regulatory peptides enhancing anxiety. Search for the optimal complexes as a basis for developing inhibitory therapeutic agents

Regulatory peptides (RP) take an active part in managing the majority of physiological processes. One of these functional applications is the monitoring of the anxiety level, of panic state. This work represents the presumptive analysis of literature data of 1960-2004 on the effects of regulatory peptides enhancing anxiety (RP-AT). This information database was used for researching the characteristics of organization and functionality of the system of anxiogenic RP. Taking into account the method of vector representation of RP effects, estimation of spectra of physiological effects that can go with each of RP-AT and their combination, was carried out. The most perspective RP complexes of anxiogenic profile for further experimental development of inhibitory therapeutic agents are proposed.     

Hagfish intestinal antimicrobial peptides are ancient cathelicidins

Three potent broad-spectrum antimicrobial peptides (HFIAP-1, -2, and -3) isolated from intestinal tissues of Myxine glutinosa (Atlantic hagfish) are identified as ancient members of the cathelicidin family of antimicrobial peptides, hitherto known only from mammals. In situ hybridization reveals that HFIAPs are produced in nests of myeloid cells within the loose connective tissue of the gut wall, a tissue reminiscent of both gut-associated lymphoid tissue (GALT) and vertebrate spleen. We suggest that this tissue organization provides local defense of the hagfish gastrointestinal tract via innate immunity and possibly served as the architectural plan upon which the adaptive immune system evolved.     

Vestibulo-protective properties of regulatory peptides

Endogenous opioid peptides take an active part in the pathogenesis of motion sickness (MS). Some regulatory peptides (substances P, gamma and des-tyr-gamma-endorphins) manifest antiemetic properties, unlike opioid peptides and morphine. Thus, we have examined the vestibulo-protective properties of some regulatory peptides during simulation of MS in cats. Vestibulo-protective properties have been discovered in some peptides: sweet water hydra undecapeptide, substances P, gamma and des-tyr-gamma-endorphins. It has been suggested that regulatory peptides take an active part in the genesis of vestibulo-protective disorders during motion sickness.     

Immunogold staining procedure for the localisation of regulatory peptides

The use of protein A- and IgG-conjugated colloidal gold staining methods for the immuno-localisation of peptide hormones and neurotransmitters at light- and electron microscope level are described and discussed. Bright-field and dark-ground illumination modes have been used to visualise the gold-labelled antigenic sites at the light microscope level. Immunogold staining procedures at the ultrastructural level using region-specific antisera have been adopted to localise specific molecular forms of peptides including gastrin (G17 and G34), glucagon and pro-glucagon, insulin and pro-insulin, in normal tissue and in tumours of the gastroenteropancreatic system. Similar methods have been used to demonstrate the heterogeneity of p-type nerves in the enteric nervous system. Vasoactive intestinal polypeptide (VIP) has been localised to granular sites (mean +/- S.D. granule diameter = 98 +/- 19 nm) in nerve terminals of the enteric plexuses and in tumour cells of diarrhoeogenic VIP-producing neoplasias (mean +/- S.D. granule diameter = 126 +/- 37 nm) using immunogold procedures applied to ultraviolet-cured ultrathin sections. Co-localisation of amines and peptides in carotid body type I cells and in chromaffin cells of normal adrenal medulla and phaeochromocytomas has also been demonstrated. Advantages of the immunogold procedures over alternative immunocytochemical techniques are discussed.     

Search for a regulatory function of mitochondrial proteinases in yeast

Proteolytic activity in yeast mitochondrial membranes and soluble fractions was studied using a labeled substrate: [¹⁴C]dimethylcasein. Optimal activity for the two fractions was at pH 6 in phosphate buffer at 37°C. Activity was partially inhibited by adding 3 mmol/1 ATP. Activity was enhanced rapidly in both fractions when yeast cells were grown aerobically on a derepression medium and transfered on a repression medium in anaerobiosis. Mitochondrial proteinase mediation in the mitochondrial breackdown observed during transition from aerobical derepression to anaerobical repression is discussed.     

Emetic and antiemetic properties of regulatory peptides

Emetic and antiemetic properties of opioid peptides, substance "P", beta-lipotropin, and ACTH1-39 have been investigated in experiments on cats. It was shown that morphine, enkephalin, beta-endorphin and DADLE caused vomiting in animals, which was blocked by naloxone. Substance "P", gamma- and des-tyr-gamma-endorphin manifested antiemetic properties similar to those of naloxone. Selective antagonists of delta-opioid receptors ICI 154, 129 blocked emetic action of delta-agonist DADLE but did not prevent vomiting caused by mu-agonist morphine. It is suggested that the vomiting mechanisms of endogenous opioid peptides involve stimulation of mu- and delta-opioid receptors in the chemoreceptor trigger zone of the vomiting centre.     

A way of functional classification of regulatory peptides. Parameters of divergent and convergent evolution of regulatory peptides

A mathematical method has been presented for systematization of functions of regulatory peptides (RP) and evaluation of directions of evolutionary development of RP systems. For this purpose, traditional methods of vector algebra and multi-dimensional space were used. Effects of various peptide regulators on anxiety, depression, and memory are considered by the example of the three-dimensional space. A way of the functional classification of peptides has been proposed.     

Sample handling techniques when analyzing regulatory peptides

Collection of blood samples in prechilled heparinized tubes, rapid cooling and centrifugation at 4 degrees C were found to be more important than the enzyme inhibitors aprotinin and EDTA in preserving immunoreactive neuropeptide Y. Nine months after storage of plasma in the frozen state at -20 degrees C or -80 degrees C the recovery of NPY was about 50% of the recovery at immediate analysis. Synthetic substance P added to guinea pig plasma at 37 degrees C disappeared almost entirely within 30 seconds as measured by radioimmunoassay while the concentrations of neurokinin A and neuropeptide K decreased only to a minor extent during a 20 min observation period. The total concentration of immunoreactive substance P and neurokinin A in boiled aqueous and acetic acid extracts of rat dorsal spinal cord was on the other hand stable for 72 h at 4 degrees C, 24 h at room temperature and after freezing and thawing three times. However, chromatographic analysis indicated that the immunoreactivity became increasingly more heterogenous in the samples particularily at room temperature. Acid ethanol and Sep Pak extraction of plasma samples resulted in almost 90% recovery of neuropeptide Y, neuropeptide K and calcitonin gene-related peptide while removing crossreacting substances with high molecular weight.     

Gut regulatory peptides in some Antarctic notothenioids

The presence and the comparative distribution of regulatory peptides and serotonin in the gut of four species of Antarctic notothenioid fishes [Cryodraco antarcticus and Chionodraco hamatus (Channichthyidae), and Trematomus bernacchii and Trematomus newnesi (Nototheniidae)], were immunohistochemically studied. In these species, numerous immunoreactive (IR) endocrine cells and nerve elements were detected. In the nototheniids most of the IR nerve fibres were of the intrinsic type, while most of the IR nerve fibres of the channichthyid intestine, besides insulin-like IR fibres, seemed to be of the extrinsic type. The intensity and frequency of immunopositivity are not the same in channnichthyids and nototheniids; the species belonging to the first family show many differences from the teleosts living in temperate water. The finding of insulin-like endocrine cells and nerve fibres in the gut wall of Cryodraco antarcticus is exceptional for vertebrates and deserves special attention.A preliminary account of this work was presented to the VII International Ichthyology Congress, Den Haag, 26–30 August 1991     

Search for partial resistance to leaf rust in a collection of ancient Spanish wheats

A collection of 917 accessions of Spanish durum and bread wheat was screened for resistance to leaf rust (Puccinia triticina) under field conditions at three locations. Resistance levels ranged from very low to very high, high susceptibility being most frequent. Relative disease severity (referred to the most susceptible accession = 100%) was lower than 20% in about 6% of the accessions in each location. In the collection most of the lines (84%) displayed a susceptible infection type. A final selection of seven accessions (one of them durum) displaying low severity level in the field and high infection type in a growth chamber was chosen for further studies. High levels of partial resistant with longer latency period and high percentage of early aborted colonies without necrosis were found. They might be used in breeding programmes.     

From peptide-bond formation to cotranslational folding: dynamic, regulatory and evolutionary aspects

Ribosomes are ribozymes exerting substrate positioning and promoting substrate-mediated catalysis. Peptide-bonds are formed within a symmetrical region, thus suggesting that ribosomes evolved by gene-fusion. Remote interactions dominate substrate positioning at stereochemistry suitable for peptide-bond formation and elaborate architectural-design guides the processivity of the reaction by rotatory motion. Nascent proteins are directed into the exit tunnel at extended conformation, complying with the tunnel's narrow entrance. Tunnel dynamics facilitate its interactive participation in elongation, discrimination, cellular signaling and nascent-protein trafficking into the chaperon-aided folding site. Conformational alterations, induced by ribosomal-recycling factor, facilitate subunit dissociation. Remarkably, although antibiotics discrimination is determined by the identity of a single nucleotide, involved also in resistance, additional nucleotides dictate antibiotics effectiveness.