Sulfoxide configuration in sparsomycin determines time-dependent and competitive inhibition of peptidyl transferase

Sparsomycin, S_cR_s configuration, was the most potent of the four possible stereoisomers as a competitive inhibitor of peptide bond formation. In addition, the configuration of the two chiral centers dictated whether the compound exhibited time- and temperature-dependent inhibition of peptidyl transferase when incubated with polysomes prior to enzyme assay. The data corroborate the thesis that a peptidyl transferase-mediated acylation of the pivotal sulfoxide moiety and subsequent Pummerer rearrangement play a significant role in the inhibitory properties of sparsomycin.     

Ultraviolet-crosslinking reveals specific affinity of eukaryotic initiation factors for Semliki Forest virus mRNA

Eukaryotic initiation factors (eIF) associate readily with ³²P-labeled Semliki Forest virus (SFV) mRNA in vitro, forming complexes which can be crosslinked by 254 nm ultraviolet irradiation. After ribonuclease digestion, the initiation factors were released and analysed by gel electrophoresis. Autoradiography revealed proteins by virtue of crosslinked ³²P-labeled mRNA fragments. eIF-4A, -4B and -4C as well as three subunits of eIF-3 could be crosslinked with SFV mRNA. None of these proteins bound to ribosomal RNAs.     

Displacement of O2 and CO by cyanide from the active site of helix pomatia β-hemocyanin.: Formation of a mixed-liganded species

Addition of KCN to Helix pomatia β-hemocyanin fully saturated with either O₂ or CO results in a decrease of the spectroscopic properties of the protein (absorbance at 340 nm and luminescence at 550 nm) due to the displacement of the gaseous ligands (O₂ or CO) from the active site. The anionic form of cyanide (CN^?) is supposed to bind to the active site; its intrinsic affinity for the protein, as calculated from independent O₂ and CO displacement experiments, is between 2 and 6 × 10⁶M^?1. The replacement of O₂ or CO shows some differences which may be correlated with the different modes of binding at the active site. Thus, while displacement of oxygen by cyanide is hyperbolic, addition of cyanide to carbonylated hemocyanin shows a lag phase. This finding suggests the formation of a mixed liganded complex at the active site. The simultaneous presence of CO and CN^? at the active site of hemocyanin is also supported by the experiment in which addition of small amounts of KCN to hemocyanin partially saturated with O₂ and CO gives rise to an increase of emission intensity and a concomitant decrease of the O₂ absorption band. The mixed-liganded species displays luminescence properties similar to those of CO-saturated hemocyanin, and the formation of the complex is reversible on dialysis or oxygenation.     

Decreased UDP-GlcNAc:Glycopeptideβ-2-N-Acetylglucosaminyltransferase II activity in a ricin-resistant mutant of baby hamster kidney (BHK) cells

The ricin-resistant mutant baby hamster kidney (BHK) cell line RIC^R21 is unable to make the sialylated bi- or triantennary complexN-glycans found in wild type cells and accumulates instead non-bisected hybrid structures containing three Man residues and one or two sialylated antennae (Hugheset al 1983, Carbohydr Res 120215-34). Specific assays forN-acetylglucosaminyltransferases I, II, III and IV were applied to Triton X-100 extracts of wild type BHK, RIC^R14 and RIC^R21 cells. It was shown that RIC^R21 cell extracts had a decreasedN-acetylglucosaminyltransferase II specific activity (17 to 27% of wild type values). It is suggested that in wild type cellsN-acetylglucosaminyltransferase II action proceeds quickly, leading to complexN-glycan synthesis, while in RIC^R21 cells potential substrates forN-acetylglucosaminyltransferase II move into the trans-Golgi compartment before the transferase can act, thereby leading to hybrid structures.     

The Winnipeg Children's Hospital child protection centre: a provincial medical initiative.

The Winnipeg Children''s Hospital child protection centre is a hospital-based provincial protection program that focuses on the diagnosis, treatment and prevention of child abuse and neglect in Manitoba. The multidisciplinary group of professionals working in the centre differs from most hospital-based teams in that it has a strong government liaison both functionally and financially, has strong ties to the welfare system and has legal counsel relating to the court system. The centre represents the philosophy that medical participation in welfare matters relating to children need not be relegated to the sidelines. This paper describes the centre''s history and structure, and it outlines some of its research projects.     

An "artificial tongue" for calibrating solution flow characteristics of taste stimulus delivery systems

An "artificial tongue" is described which can be used to calibratethe stimulus delivery characteristics of flow systems for liquidtaste stimuli. The calibrator, which functions on the basisof conductance measurements, is used to ascertain the behaviourof certain test solutions in the flow system. The test solutionsare so prepared that they mimic the hydraulic properties ofthe stimulus solutions to be used. The special apparatus requiredis neither complex nor costly, and the procedures involved arestraightforward.     

Resolution of optical isomers by high-pressure liquid chromatography: The separation of benzo[a]pyrene trans-diol derivatives

The optical isomers of (±)r-7,t-8-dihydroxy-7,8-dihydrobenzo[a]pyrene and its synthetic precursor (±)r-7,t-8-dihydroxy-7,8,9,10-tetrahydrobenzo[a]pyrene were resolved as their di-(−)menthoxyacetates using high-pressure liquid chromatography. Saponification of the resolved diesters yielded the corresponding enantiomers. The specific rotation, CD spectra, and ORD curves are reported. The resolution of these optical isomers permits detailed studies on the enzymatic intermediates and the mechanism of benzo[a]pyrene activation to its carcinogenic form. The method is of general usefulness for the resolution of optical isomers.     

High-resolution nuclear magnetic resonance determination of transfer RNA tertiary base pairs in solution. 1. Species containing a small variable loop.

Eight class I tRNA species have been purified to homogeneity and their proton nuclear magnetic resonance (NMR) spectra in the low-field region (-11 to -15 ppm) have been studied at 360 MHz. The low-field spectra contain only one low-field resonance from each base pair (the ring NH hydrogen bond) and hence directly monitor the number of long-lived secondary and tertiary base pairs in solution. The tRNA species were chosen on the basis of their sequence homology with yeast phenylalanine tRNA in the regions which form tertiary base pairs in the crystal structure of this tRNA. All of the spectra show 26 or 27 low-field resonances approximately 7 of which are derived from tertiary base pairs. These results are contrary to previous claims that the NMR spectra indicate the presence of resonances from secondary base pairs only, as well as more recent claims of only 1-3 tertiary resonances, but are in good agreement with the number of tertiary base pairs expected in solution based on the crystal structure. The tertiary base pair resonances are stable up to at least 46 degrees C. Removal of magnesium ions causes structural changes in the tRNA but does not result in the loss of any secondary or tertiary base pairs.     

Evolutionary temperature adaptation of fish sarcoplasmic reticulum

Summary Sarcoplasmic reticulum has been isolated from the white muscle of 15 species of teleost fish adapted to diverse thermal environments. Evidence has been obtained that the Ca²⁺-dependent ATPase of fish sarcoplasmic reticulum has undergone evolutionary modification for function at different temperatures. Compared with tropical fish, cold adapted species have higher rates of Ca²⁺ transport and Ca²⁺-ATPase activities at low temperatures. Most species have linear Arrhenius plots over the temperature range 0–30°C. Activation enthalpies (H ) of the ATPase ranged from 53–190 kJ mol^–1 and were positively correlated with environment temperature. Activation entropy (S ) varied from negative values in cold adapted species to positive values in tropical fish.In contrast to the Ca²⁺-ATPase, the basal ATPase of fish sarcoplasmic reticulum showed no relationship between either ATPase activity or thermodynamic activation parameters and environmental temperature.Only the Ca²⁺-dependent ATPase is coupled to Ca²⁺ transport. The percentage of total ATPase activity which is Ca²⁺ activated is higher at low temperatures in cold than in warm adapted species. For example, ratios of Ca²⁺-dependent/total ATPase at 2°C varied from 80–98% in Arctic, Antarctic and North Sea species to only 2–50% in various tropical fish. Above 20°C, similar ratios in the range 80–98% were obtained for all species. The nature of the basal ATPase and mechanisms of temperature adaptation of fish sarcoplasmic reticulum are discussed.Abbreviations ET environmental temperature - EGTA ethylene glycol-bis (-aminolethyl ether)-N, N-tetraacetic acid - HEPES N-2-hydroxylpiperazine-N-2-ethanesulfonic acid - SR sarcoplasmic reticulum