Neo-kyotorphin (Thr-Ser-Lys-Tyr-Arg), a new analgesic peptide

The amino acid sequence of a newly isolated pentapeptide, neo-kyotorphin from bovine brain was synthetically verified to be Thr-Ser-Lys-Tyr-Arg corresponding to the C-terminal portion of hemoglobin alpha-chain. The synthetic neo-kyotorphin showed the dose-dependent analgesia in mice which was approximately equal to that of Leu-enkephalin.     

Pituitary adenylate cyclase-activating peptide (PACAP), a new vasoactive intestinal peptide (VIP)-like peptide in the respiratory tract

Summary Pituitary adenylate cyclase-activating peptide (PACAP) is a vasoactive intestinal peptide (VIP)-like peptide recently isolated from ovine hypothalami. Nerve fibers displaying PACAP immunoreactivity were found in the respiratory tract of rats, guinea pigs, ferrets, pigs, sheep and squirrel monkeys. A moderate supply of PACAP-immunoreactive fibers was seen in the nasal mucosa of guinea pigs. Few to moderate numbers of PACAP-containing fibers occurred in the tracheo-bronchial wall of rats, guinea pigs, ferrets, pigs, sheep and squirrel monkeys. The fibers were distributed beneath the epithelium, around blood vessels and seromucous glands, and among bundles of smooth muscle. In the lungs, the immunoreactive fibers were observed close to small bronchioli. A few PACAP-immunoreactive nerve cell bodies were seen in the sphenopalatine and otic ganglia of guinea pigs. Simultaneous double immunostaining of the respiratory tract of sheep and ferrets revealed that all PACAP-containing nerve fibers stored VIP. We suggest that neuronal PACAP may take part in the regulation of smooth muscle tone and glandular secretion.     

Ontogeny of a new enteric peptide, neuropeptide W (NPW), in the developing rat stomach

Neuropeptide W (NPW), a novel endogenous peptide for G protein-coupled receptors GPR7 and GPR8, is expressed in the gastric antral mucosa of rat, mouse, and human stomachs. Here, we studied the ontogeny of NPW in the developing rat stomach. Real-time RT-PCR showed that NPW gene expression was initially detectable in embryonic day 14 (E14) stomach and gradually increased during the progress of age until birth, postnatal day 1 (P1). NPW mRNA level in the stomach increased again from the weaning period (P21) until reaching adulthood. Immunohistochemistry using polyclonal antibodies raised against rat NPW revealed that NPW-positive cells were detected in the P1 antral stomach and gradually increased during the development of age. Furthermore, double immunohistochemistry demonstrated that NPW colocalized with gastrin in P1 rat stomach. These data will provide clues to physiological functions of NPW in the development of rat stomach.     

Radioimmunoassay for a new phenothiazine derivative and its application.

Fluphenazine-4-chlorophenoxy-isobutyrate ester, a new phenothiazine derivative was synthesized in the Institute for Drug Research Budapest. Radioimmunoassay was developed for the therapeutic monitoring of the drug level after intramuscular depot injection. The fluphenazine hapten was coupled to BSA by mixed-anhydride method. Antisera were produced to this conjugation in New-Zealand white rabbits and were tested for the antibody-titer. The specificity was tested by the cross-reaction with phenothiazine-analogues and other psychotropics. Strong cross-reaction was found with compounds possessing piperazine in side chain (trifluoperazine, perphenazine), but other psychotropic drugs did not react. Tritium-labelled trifluoperazine (spec. activity: 3.5 TBq/mmol) was used as a tracer in the radioimmunoassay. The detection limit was 75 pg with a CV of < 5% in 50 microliters plasma sample (equivalent to 1.5 ng/ml concentration) and a standard curve in the 3 ng/ml-50 ng/ml GYKI-22441 concentration range showed a CV of < 10%. Preliminary pharmacokinetic study was performed in Beagle dogs after intramuscular depot injection with GYKI-22441 in sesame oil in a dose of 0.1 mg/kg. The GYKI-22441 concentration of the plasma samples were measured by the RIA method during a 28-day interval after the treatment and was evaluated by the MultiCalc Immunoassay Data Management program (Pharmacia).     

A new amino acid derivative with a masked side-chain aldehyde and its use in peptide synthesis and chemoselective ligation.

A new amino acid derivative with a diol side-chain, L-2-amino-4,5-dihydroxy-pentanoic acid (Adi), has been prepared from L-allylglycine by suitable protection, for use in peptide synthesis, as Fmoc-L-Adi(Trt)2. This building block enables the introduction of a side-chain aldehyde at any position in a given peptide sequence without use of specialized side-chain protection schemes. The aldehyde is revealed by mild oxidation with sodium periodate, circumventing the problematic release of reactive peptidic aldehydes in TFA solution. Peptides with aldehyde side-chains are useful for chemo-selective ligation, reacting selectively with oxyamines to yield oxime links, while all other peptide functions can be left unprotected. The utility of the new building block has been demonstrated by the synthesis of peptide dimers and a cyclo-peptide.     

The development of the eye in Astyanax mexicanus (Characidae,Pisces), its blind cave derivative,Anoptichthys Jordani (Characidae,Pisces), and their crossbreds

The development of the eye of the characin Astyanax mexicanus, of its blind derivative Anoptichthys jordani, and crossbreds of both forms was studied at different ontogenetic stages by means of scanning- and transmission-electron microscopy. Astyanax exhibits a form of eye development resembling that in other characid species. A severe reduction of the eye could be observed in Anoptichthys starting with the second day of ontogeny. This degenerational process is characterized by the following features: 1) An overgrowth of epidermal tissue that gradually covers the surface of the eyeball; 2) the sinking of the eyeball below the surface of the integument; 3) the formation of epidermal channels from the body surface to the disappearing surface of the eyeball; 4) a severe degeneration of the retinal sensory cells; and 5) a small number of pigment granules in the pigment epithelial cells. The progeny of crosses between Astyanax and Anoptichthys show varying degrees of these degenerational signs. Taste buds and the lateral line organ display identical features in all crosses analyzed with the scanning electron microscope.     

Solution structures and biological functions of the antimicrobial peptide, arenicin-1, and its linear derivative

Arenicin-1 (AR-1) is a novel antimicrobial peptide that was isolated from coelomocytes of the marine polychaeta lugworm Arenicola marina and shown to contain a single disulfide bond between Cys3 and Cys20, forming an 18-residue ring [Ovchinnikova, T. V. et al., FEBS Lett 2004, 577, 209-214]. To determine the role of this disulfide bond, we synthesized AR-1 (RWCVYAYVRVRGVLVRYRRCW) and its linear derivative, arenicin-1-S (AR-1-S: RWSVYAYVRVRGVLVRYRRSW). Activity assays revealed that AR-1-S is somewhat less active against bacterial cells than AR-1. Both peptides were very hydrophobic, and displayed cytotoxicity against human red blood cells. Analysis of the tertiary structures of AR-1 and AR-1-S by NMR spectroscopy disclosed that AR-1 has two-stranded antiparallel beta-sheet structures with amphipathicity, while AR-1-S displays a random coil structure in DMSO. Our biological data for AR-1 and AR-1-S indicate that the hydrophobic-hydrophilic balance, disulfide bridge, and the amphipathic beta-sheet structure of the peptides play important roles in their biological activities. Elucidation of the structure of AR-1 and its derivative should facilitate the design of novel non-cytotoxic peptide antibiotics with potent antibacterial activities.     

Kinetic and thermodynamic analyses of adhesion of a peptide,Trp-Lys-Tyr-Met-Val-D-Met (WKYMVm), and human formyl peptide receptor (hFPR)

The kinetic and thermodynamic properties of a peptide–receptor interaction was investigated by measuring the adhesion force in the reaction via atomic force microscopy (AFM). Trp-Lys-Tyr-Met-Val-D-Met (WKYMVm), considered as a model system in the present study, is a potent neutrophil chemo-attractant. Since being identified as an agonist for formyl peptide receptor (FPR), WKYMVm’s high affinity to FPR has been verified through investigation of its kinetic and physiological behaviors via conventional methods. However, there have been no reports on the adhesion force of WKYMVm-FPR. In this research, we measured the adhesion force of WKYMVm-FPR using AFM. Kinetic parameters obtained from the relationship between the adhesion force and loading rate were used to characterize the thermodynamic properties of WKYMVm-hFPR binding.     

Synthesis, crystal structures, and molecular hyperpolarizabilities of a new Schiff base ligand, and its copper(II), nickel(II), and cobalt(II) metal complexes

A new ligand (HL) obtained from the Schiff base condensation of 4-(diethylamino)salicylaldehyde with 4-nitroaniline is reported, with its nickel(II), copper(II), and cobalt(II) complexes. The crystal structures are reported for the four derivatives. While, Ni^IIL₂ and Cu^IIL₂ are centrosymmetric molecules, Co^IIL₂ exhibits a pseudo-tetrahedral molecular structure. The quadratic hyperpolarizabilities (β) of HL and Co^IIL₂, measured by electric field induced second harmonic (EFISH) technique, are equal to 66 and 110 × 10⁻³⁰ cm⁵ esu⁻¹, respectively. Beside a geometric effect (pseudo-T_d symmetry), the coordination of the metal center provides an intrinsic enhancement of the NLO response. In addition, an enhancement of the thermal stability of about 60° is found upon metal complexation.     

Anti-HIV-1 activity of a new scorpion venom peptide derivative Kn2-7

For over 30 years, HIV/AIDS has wreaked havoc in the world. In the absence of an effective vaccine for HIV, development of new anti-HIV agents is urgently needed. We previously identified the antiviral activities of the scorpion-venom-peptide-derived mucroporin-M1 for three RNA viruses (measles viruses, SARS-CoV, and H5N1). In this investigation, a panel of scorpion venom peptides and their derivatives were designed and chosen for assessment of their anti-HIV activities. A new scorpion venom peptide derivative Kn2-7 was identified as the most potent anti-HIV-1 peptide by screening assays with an EC(50) value of 2.76 μg/ml (1.65 μM) and showed low cytotoxicity to host cells with a selective index (SI) of 13.93. Kn2-7 could inhibit all members of a standard reference panel of HIV-1 subtype B pseudotyped virus (PV) with CCR5-tropic and CXCR4-tropic NL4-3 PV strain. Furthermore, it also inhibited a CXCR4-tropic replication-competent strain of HIV-1 subtype B virus. Binding assay of Kn2-7 to HIV-1 PV by Octet Red system suggested the anti-HIV-1 activity was correlated with a direct interaction between Kn2-7 and HIV-1 envelope. These results demonstrated that peptide Kn2-7 could inhibit HIV-1 by direct interaction with viral particle and may become a promising candidate compound for further development of microbicide against HIV-1.     

Synthesis and properties of cholecystokinin-releasing peptide (monitor peptide), a 61-residue trypsin inhibitor

A 61-residue cholecystokinin-releasing peptide (monitor peptide), which was obtained from rat pancreatic juice and found to stimulate pancreatic enzyme secretion, was recently reported to inhibit bovine trypsin and to possess epidermal growth factor (EGF)-like activities, at a concentration of about 10 nM. However, monitor peptide is structurally different from the EGF family of growth factors. To investigate whether monitor peptide contains the supposed EGF-like activities, it has been synthesized together with its [Ala23, Ala47] analog. The purified peptides, which were fully characterized by a range of methods including Cf-252 ionization mass spectrometry and enzymatic digestion to establish the locations of disulfide linkages, were shown to belong to the pancreatic secretory trypsin inhibitor family and not to the EGF family. Neither synthetic monitor peptide nor its analog were able to compete with 125I-EGF in A-431 cells or to stimulate growth of Swiss 3T3 and NRK 49F cells, up to 1 microM concentration. However, synthetic monitor peptide was as effective as the native product in the inhibition of trypsin. Replacement of the essential Arg23 in the [Ala23, Ala47]-analog led to loss of trypsin inhibition activity.     

Biotransformation of a 4(20),11(12)-taxadiene derivative

A 4(20),11(12)-taxadiene derivative was converted to hydroxylated derivatives by Cunninghamella elegans AS3.2033 and Cunninghamella elegans var chibaensis ATCC 20230. Both microorganisms led to C-1 hydroxylations and conversion to a C-15-hydroxylated abeo-taxane. Additional products from the two fungi differed: a C-14 oxidation and a trans-cis isomerization of the cinnamoyl for one and an unprecedented hydroxylation at C-17 for the other.     

Lactoferricin, a new antimicrobial peptide

Lactoferricin B (LF-B) is a peptide derived from acid-pepsin digestion of bovine lactoferrin, which has antimicrobial properties. In order to assess the antimicrobial spectrum of LF-B and its possible in vivo uses, the minimum inhibitory and microbicidal concentrations of pure lactoferricin B were determined for a range of bacterial species and under varying conditions of growth including growth phase and size of the inoculum, pH and ionic strength of the medium. Lactoferricin B was bactericidal against a wide range of bacteria and Candida albicans. Proteus spp., Pseudomonas cepacia and Serratia spp. were resistant. The bactericidal activity of LF-B was inhibited by increasing ionic strength and bacterial inoculum and at acid pH. The activity of lactoferricin B was completely inhibited by the addition of 5% whole cow's milk and was reduced in the presence of increasing concentrations of mucin. These results indicate the potential of LF-B to reduce the numbers of organisms in a simple medium, but raise doubts about its role in vivo because of its sensitivity to changes in physical variables. It may be that lactoferricin exerts a transient antimicrobial effect at mucosal surfaces.     

Deconica cokeriana (Agaricales,Strophariaceae), a new combination

Deconica cokeriana is proposed as a new combination based on morphological and molecular data. This species is fully described, discussed, and illustrated. Until now, D. cokeriana is only known from the eastern USA, specifically from Connecticut, Georgia, Ohio, and Tennessee.     

Crystallization of a derivative of a new coenzyme,methoxatin

A new compound, derived from a parent compound to which we have given the trivial name, methoxatin, has been isolated from a methanol-oxidizing bacterium, and crystallized. Its chemical structure was determined by X-ray crystallography. Methoxatin is implicated as a coenzyme in the oxidation of substrate alcohols. This report describes the purification and crystallization of the derivative, acetonyl methoxatin.