Sense and antisense peptides, i.e. peptides specified by complementary DNA and RNA sequences, interact with increased probability. Biro, Blalock, Mekler, Root-Bernstein and Siemion investigated the recognition rules of peptide-peptide interaction based on the complementary coding of DNA and RNA sequences in 3'→5' and 5'→3' directions. After more than three decades of theoretical and experimental investigations, the efficiency of this approach to predict peptide-peptide binding has been experimentally verified for more than 50 ligand-receptor systems, and represents a promising field of research. The natural genetic coding algorithm for sense and antisense peptide interactions combines following elements: of amino acid physico-chemical properties, stereochemical interaction, and bidirectional transcription. The interplay of these factors influences the specificity of sense-antisense peptide interactions, and affects the selection and evolution of peptide ligand-receptor systems. Complementary mRNA codon-tRNA anticodon complexes, and recently discovered Carter-Wolfenden tRNA acceptor-stem code, provide the basis for the rational modeling of peptide interactions based on their hydrophobic and lipophilic amino acid physico-chemical properties. It is shown that the interactions of complementary amino acid pairs according to the hydrophobic and lipophilic properties strongly depend on the central (second) purine base of the mRNA codon and its pyrimidine complement of the tRNA anticodon. This enables the development of new algorithms for the analysis of structure, function and evolution of protein and nucleotide sequences that take into account the residue's tendency to leave water and enter a nonpolar condensed phase considering its mass, size and accessible surface area. The practical applications of the sense-antisense peptide modeling are illustrated using different interaction assay types based on: microscale thermophoresis (MST), tryptophan fluorescence spectroscopy (TFS), nuclear magnetic resonance spectroscopy (NMR), and magnetic particles enzyme immunoassay (MPEIA). Various binding events and circumstances were considered, e.g., in situations with-short antisense peptide ligand (MST), L- and D-enantiomer acceptors (TFS), in low affinity conditions (NMR), and with more than one antisense peptide targeting hormone (MPEIA).

Antisense peptide technology is a valuable tool for deriving new biologically active molecules and performing peptide–receptor modulation. It is based on the fact that peptides specified by the complementary (antisense) nucleotide sequences often bind to each other with a higher specificity and efficacy. We tested the validity of this concept on the example of human erythropoietin, a well-characterized and pharmacologically relevant hematopoietic growth factor. The purpose of the work was to present and test simple and efficient three-step procedure for the design of an antisense peptide targeting receptor-binding site of human erythropoietin. Firstly, we selected the carboxyl-terminal receptor binding region of the molecule (epitope) as a template for the antisense peptide modeling; Secondly, we designed an antisense peptide using mRNA transcription of the epitope sequence in the 3'→5' direction and computational screening of potential paratope structures with BLAST; Thirdly, we evaluated sense–antisense (epitope–paratope) peptide binding and affinity by means of fluorescence spectroscopy and microscale thermophoresis. Both methods showed similar Kd values of 850 and 816 µM, respectively. The advantages of the methods were: fast screening with a small quantity of the sample needed, and measurements done within the range of physicochemical parameters resembling physiological conditions. Antisense peptides targeting specific erythropoietin region(s) could be used for the development of new immunochemical methods. Selected antisense peptides with optimal affinity are potential lead compounds for the development of novel diagnostic substances, biopharmaceuticals and vaccines.

Iron transport across the periplasmic space to the cytoplasmic membrane of certain Gram-negative bacteria is mediated by a ferric binding protein (Fbp). This requires Fe(3+) loading of Fbp at the inner leaflet of the outer membrane. A synergistic anion is required for tight Fe(3+) sequestration by Fbp. Although phosphate fills this role in the protein isolated from bacterial cell lysates, nitrilotriacetate anion (NTA) can also satisfy this requirement in vitro. Here, we report the kinetics and mechanism of Fe(3+) loading of Fbp from Fe(NTA)(aq) in the presence of phosphate at pH 6.5. The reaction proceeds in four kinetically distinguishable steps to produce Fe(3+)Fbp(PO(4)) as a final product. The first three steps exhibit half-lives ranging from ca. 20 ms to 0.5 min, depending on the concentrations, and produce Fe(3+)Fbp(NTA) as an intermediate product of significant stability. The rate for the first step is accelerated with an increasing phosphate concentration, while that of the third step is retarded by phosphate. Conversion of Fe(3+)Fbp(NTA) to Fe(3+)Fbp(PO(4)) in the fourth step is a slow process (half-life approximately 2 h) and is facilitated by free phosphate. A mechanism for the Fe(3+)-loading process is proposed in which the synergistic anions, phosphate and NTA, play key roles. These data suggest that not only is a synergistic anion required for tight Fe(3+) sequestration by Fbp, but also the synergistic anion plays a critical role in the process of inserting Fe(3+) into the Fbp binding site.

Abstract. Kinetics and mechanisms of the oxidation of methoxyurea and N-methylhydroxyurea were studied in neutral and basic aqueous solutions. The obtained pH dependences of the oxidation rates indicate that for both hydroxyureas the reactive species are the deprotonated ones. The second order rate constants, the activation enthalpies and the activation entropies for the reactions of methoxyurea (O-methylhydroxyurea) and N-methylhydroxyurea anions with Fe(CN) 6 3− at 25 o C, I = 2 mol dm −3 (NaClO 4 ) were determined as (5.06 ± 0.01)  10 2 mol −1 dm 3 s −1 , (1.92 ± 0.02)  10 4 mol −1 dm 3 s −1 , 27 ± 1 kJ mol, and 107 ± 4 J mol −1 K −1 , respectively. The pK a value of methoxyurea at 25 o C and 2 mol dm −3 ionic strength was determined kinetically as 12.7 ± 0.1 and the thermodynamic parameters for the deprotonation reaction were determined as Δ a H = 43 ± 1 kJ mol, and. When the kinetic results are compared with the data reported for hydroxyurea, an inverse dependence of the rate constants on the pK a of the hydroxyurea derivatives at 25 o C is observed. Such unexpected behaviour has been explained by the ab initio calculations and NBO analysis of HOMOs for all three hydroxyureates. (doi: 10.5562/cca1799)