Evolutionary conservation of the chromosomal configuration and regulation of amylase genes among eight species of the Drosophila melanogaster species subgroup

Nuclear DNA was extracted from each of the eight species comprising the Drosophila melanogaster species subgroup. Southern hybridization of this DNA by using a molecular probe specific for the alpha-amylase coding region showed that the duplicated structure of the amylase locus, first found in D. melanogaster, is conserved among all species of the melanogaster subgroup. Evidence is also presented for the concerted evolution of the duplicated genes within each species. In addition, it is shown that the glucose repression of amylase gene expression, which has been extensively studied in D. melanogaster, is not confined to this species but occurs in all eight members of the species subgroup. Thus, both the duplicated gene structure and the glucose repression of Drosophila amylase gene activity are stable over extended periods of evolutionary time.      

Binding and action of cecropin and cecropin analogues: antibacterial peptides from insects

The mechanism of action of cecropin was studied by using liposomes as a model system. The bilayer was efficiently destroyed if the liposome net charge was zero or negative. Cecropin analogues with an impaired N-terminal helix had reduced membrane disrupting abilities that correlate with their lower antibacterial activity. The reduced bactericidal activity of the analogues was rationalized in terms of reduced binding to bacteria. The stoichiometry of cecropin killing of bacteria suggests that amounts of cecropin sufficient to form a monolayer strongly modify the bacterial membrane. Although some bacteria were resistant to cecropin they did bind large amounts in a non-productive manner. In contrast, mammalian erythrocytes achieve resistance by avoiding the binding of cecropin.     

Molecular basis of superreactivity of cysteine residues 31 and 32 of seminal ribonuclease

The molecular basis of the high reactivity toward reducing agents of intersubunit disulfides at positions 31 and 32 of dimeric bovine seminal ribonuclease was investigated by studying in the monomeric enzyme the fast reaction kinetics with disulfides of the adjacent cysteine-31 and -32, exposed by selective reduction of the intersubunit disulfides. Negatively charged and neutral disulfide reagents were used for measuring the thiol reaction rates at neutral pH. The kinetics studied as a function of pH permitted us to define pK values for the thiols of interest and indicated the possibility of determining pK values of SH groups in proteins indirectly by measuring the kinetics of reactivity of the SH groups with a disulfide reagent. The results were compared with those obtained under identical conditions with synthetic thiol peptides and model compounds. The data indicate that the superreactivity of intersubunit disulfides of seminal ribonuclease is matched by the high reactivity at neutral pH of adjacent cysteine residues 31 and 32, as compared to all small thiol compounds tested. The synthetic hexapeptide segment of seminal ribonuclease Ac-Met-Cys-Cys-Arg-Lys-Met-OH, which includes the two cysteine residues of interest, was even more reactive. These data, and the other results reported in this paper, led to the conclusion that the superreactivity at neutral pH of cysteine residues at positions 31 and 32 of bovine seminal ribonuclease is primarily dependent on the nearby presence of positively charged groups, particularly the epsilon-NH2 of lysine-34, and is influenced by the adjacency of the two thiols and by the protein tertiary structure.     

The reaction of alkylating agents with bacteriophage R17. Biological effects of phosphotriester formation

The extent of biological inactivation and of the degradation of the RNA after reaction of bacteriophage R17 with ethyl methanesulphonate, isopropyl methanesulphonate and N-ethyl-N-nitrosourea was studied. Formation of breaks in the RNA chain probably results from hydrolysis of phosphotriesters formed in the alkylation reactions. Near neutral pH the ethyl and isopropyl phosphotriesters are sufficiently stable for the kinetics of the hydrolysis reaction to be followed. Results indicate that the rate of hydrolysis increases rapidly as the pH is raised. The evidence shows that a phosphotriester group does not itself constitute a lethal lesion. The extent of phosphotriester formation by the different agents is discussed in terms of reaction mechanism.     

Vitamin K5 as a Fungistatic Agent

The effectiveness of vitamin K(5) in controlling the growth of different molds at varying pH levels in a culture medium, in tomato juice, and in several berry purees was studied. The molds studied were Aspergillus, Botrytis, Hormodendrum, Mucor, and Penicillium. The results showed that vitamin K(5) was effective as a fungistatic agent at concentrations ranging from 0.006 to 0.02%.     

Smith degradation of gum exudates from some prosopis species

The polysaccharide of P. hymantophora has been shown to be composed of (1→4)-linked galactopyranosyl, (1→3)-linked galactopyranosyl, (1→3)-linked galactopyranosyl 2- and 4-sulphate and 2,6-disulphate residues. The (1→3)- and (1→4)-linked units are present in approximately equal amounts. The polysaccharide of P. hieroglyphica has been shown to possess (1→4)-linked galactopyranosyl, (1→3)-linked galactopyranosyl, and (1→3)-linked galactopyranosyl 2- and 4-sulphate residues. The (1→3)- and (1→4)-linked units are present in a 4:1 ratio. Both polysaccharides contain small proportions of non-reducing xylosyl end-groups.     

A covalently bound photoisomerizable agonist. Comparison with reversibly bound agonists at electrophorus electroplaques

After disulphide bonds are reduced with dithiothreitol, trans-3- (α-bromomethyl)-3’-[α- (trimethylammonium)methyl]azobenzene (trans-QBr) alkylates a sulfhydryl group on receptors. The membrane conductance induced by this “tethered agonist” shares many properties with that induced by reversible agonists. Equilibrium conductance increases as the membrane potential is made more negative; the voltage sensitivity resembles that seen with 50 [mu]M carbachol. Voltage- jump relaxations follow an exponential time-course; the rate constants are about twice as large as those seen with 50 μM carbachol and have the same voltage and temperature sensitivity. With reversible agonists, the rate of channel opening increases with the frequency of agonist-receptor collisions: with tethered trans-Qbr, this rate depends only on intramolecular events. In comparison to the conductance induced by reversible agonists, the QBr-induced conductance is at least 10-fold less sensitive to competitive blockade by tubocurarine and roughly as sensitive to “open-channel blockade” bu QX-222. Light-flash experiments with tethered QBr resemble those with the reversible photoisomerizable agonist, 3,3’,bis-[α-(trimethylammonium)methyl]azobenzene (Bis-Q): the conductance is increased by cis {arrow} trans photoisomerizations and decreased by trans {arrow} cis photoisomerizations. As with Bis-Q, ligh-flash relaxations have the same rate constant as voltage-jump relaxations. Receptors with tethered trans isomer. By comparing the agonist-induced conductance with the cis/tans ratio, we conclude that each channel’s activation is determined by the configuration of a single tethered QBr molecule. The QBr-induced conductance shows slow decreases (time constant, several hundred milliseconds), which can be partially reversed by flashes. The similarities suggest that the same rate-limiting step governs the opening and closing of channels for both reversible and tethered agonists. Therefore, this step is probably not the initial encounter between agonist and receptor molecules.