Subcellular organization of Streptomyces aureofaciens and overproduction of chlortetracycline

Biosynthesis of chlortetracycline is localized differently under low- and high-production conditions (standard low-production strain and its high-production variant). The experimental evidence was based on the assay of anhydrotetracycline oxygenase in subcellular fractions, ultracytochemical localization and electron-probe X-ray microanalysis of the product in the mycelium. Overproduction of chlortetracycline is closely associated with compartmentation of biosynthetic enzymes and with an efficient export of the antibiotic out of the cell.     

Timing of laparoscopic aspiration of preovulatory oocytes in heifers

Twenty-one heifers were synchronized with PGF(2) alpha and 22 heifers were stimulated with FSH-P in decreasing doses and synchronized with PGF(2) alpha. The beginning of LH rise was observed to be 47.6+/-14.0 h and 35.9+/-1.3 h (P < 0.05) and the peak of LH rise was observed at 53.8 +/- 13.4 h and 40.14+/-1.4 h (P < 0.05) after the luteolyticum administration in the synchronized and superovulated group respectively. The beginning of LH rise was observed 6.9+/-6.7 h and 4.8+/-2.6 h (P < 0.05) and the LH peak was observed 11.8+/-7.7 h and 8.3+/-3.2 h (P < 0.05) after the frist symptoms of oestrus in the synchronized and superovulated group respectively. Ovulation was not observed in stimulated heifers in the period of 23-25 h after the preovulatory LH rise. Compact cumulus oophorus was seen at 30.5%, expanded at 67.0 and partial at 2.5% during this interval of 23-25h. Within this same interval 26.9%, 51.3% and 21.8% oocytes without perivitelline space, with perivitelline space and with extruded first polar body were aspirated respectively. It may be concluded from the reported results that to recover fully mature oocytes for in vitro fertilization, it will be necessary to monitor the preovulatory period of the donor cow in great detail.     

Phospholipids chiral at phosphorus: Fourier-transform infrared study on the gel-liquid crystalline transition of chiral thiophosphatidylcholine

Fourier-transform infrared spectroscopy (FT-IR) was used to study the structural properties of Rp, Sp, and Rp + Sp isomers of 1,2-dipalmitoyl-sn-glycero-3-thiophosphocholine (DPPsC), in comparison with those of 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC). For the vibrational modes of acyl chains, isomers of DPPsC show similar temperature and phase dependence to DPPC. However, the Rp isomer of DPPsC exhibits several unique properties: the CH2 symmetric stretching band is unusually weak, the CH2 asymmetric stretching band is unusually narrow, and the CH2 wagging bands do not disappear completely at temperatures above the main transition. These differences could imply a tighter packing and be responsible for the unique phase-transition property of (Rp)-DPPsC. For the vibrational modes of the thiophosphodiester group, the frequency of the P-O stretching mode of DPPsC suggests that the POS- triad exists predominantly in the mesomeric form. This is in contrast to the structure of nucleoside phosphorothioates where charge localization at sulfur has been demonstrated [Iyengar, R., Eckstein, F., & Frey, P. A. (1984) J. Am. Chem. Soc. 106, 8309-8310]. This suggests that the different biophysical properties between isomers of DPPsC are not due to different charge distribution in the POS- triad or different geometry of charge distribution on the membrane surface. Instead, factors such as size or hydration property of oxygen and sulfur, as well as the different configuration at phosphorus, could be responsible for the differences in the conformation and packing of acyl chains, as revealed by the different properties in the CH2 stretching and wagging modes of DPPsC.     

Alpha and beta forms of cytochrome c oxidase observed in rat heart myocytes by low temperature Fourier transform infrared spectroscopy

Carbon monoxide bound to myoglobin and cytochrome c oxidase in separated adult rat heart myocytes has been observed with Fourier transform IR spectroscopy at low temperatures. CO complexes of these two proteins can be spectrally separated through temperature manipulation of the relaxation of the photolyzed systems. Photolyzed carboxymyoglobin relaxes very rapidly above 80 K, whereas the CO photolyzed from cytochrome a3 associates with CuB and relaxes very slowly below 140 K. Cytochrome c oxidase is found to be present in two major molecular forms which we designate alpha and beta. Each form contains an a3Fe and its associated CuB which we observe by their CO complexes. The predominant FeCO band, the alpha form of cytochrome oxidase, is similar to that previously seen in beef heart mitochondria, but with a slightly larger activation enthalpy, delta H = 46 kJ/mol. At least one of the beta forms is similar, but two have not been observed in beef heart mitochondria. Upon photolysis of alpha-FeCO, the alpha-CuCO species is formed. This band splits into two at low temperature. Up to half of the FeCO band area of the intact myocytes is distributed among three or more minor species (beta forms). The beta-FeCO bands all appear to be associated with only one beta-CuCO band which does not split at low temperature. After photo-dissociation of CO, the beta forms relax considerably faster than the alpha form, achieving 50% recombination in 10% of the time required for the alpha form. In a tissue slice from an opossum heart exposed to CO, we observed alpha and beta forms of cytochrome oxidase very similar to those in the rat heart myocytes. The cause of the differences between the alpha and beta forms of the enzyme is unknown, but their possible role in the control of respiration is discussed. Carboxymyoglobin contained within intact rat heart myocytes was very similar to sperm whale carboxymyoglobin, but with a much smaller amount of the lower frequency minor component.     

An improved method to isolate lichen algae by gel filtration

Photobiont cells of the lichen Evernia prunastri have completely been separated from their fungal partner by filtration through a bed of Sepharose 2B. Both mannitol and ribitol have been quantified by gas-liquid chromatography in the different steps of the isolation procedure. Absence of mannitol, which is exclusively produced by the mycobiont, has been used as the best probe to monitor isolation.     

Invertase immobilization via its carbohydrate moiety

After periodate oxidation of its glycosidic component, invertase was covalently bound onto three types of modified solid supports: glycidyl methacrylate, styrene-divinylbenzene copolymers, and bead cellulose. Direct reaction of the invertase aldehyde groups that were formed with amino groups of the support and use of the modified Ugi reaction have been employed as immobilization procedures. Apart from binding methods, the important effects of the buffer, support, conditions of periodate oxidation, and the length of the spacer on the activity of the enzyme conjugate have been investigated. Superior conjugate activity was obtained, via modified Ugi reaction, by the immobilization of a suitably oxidized invertase to a styrene-divinylbenzene copolymer having free amino groups.     

Base pairing in wheat germ ribosomal 5S RNA as measured by ultraviolet absorption, circular dichroism, and Fourier-transform infrared spectrometry

Ultraviolet absorption (UV) and circular dichroism (CD) spectra of wheat germ 5S RNA, when compared to tRNAPhe, indicate a largely base-paired and base-stacked helical structure, containing up to 36 base pairs. Fourier-transform infrared (FT-IR) spectra of tRNAPhe and wheat germ ribosomal 5S RNA have been acquired at 30 and 90 degrees C. From the difference of the FT-IR spectra between 90 and 30 degrees C, the number of base pairs in both RNAs was determined by modification of a previously published procedure [Burkey, K. O., Marshall, A. G., & Alben, J. O. (1983) Biochemistry 22, 4223-4229]. The base-pair composition and total base-pair number from FT-IR data are now consistent for the first time with optical (UV, CD, Raman) and NMR results for ribosomal 5S RNA. Without added Mg2+, tRNAPhe gave 18 +/- 2 base pairs [7 A-U and 11 G-C], in good agreement with the number of secondary base pairs from X-ray crystallography [8 A-U, 12 G-C, and 1 G-U]. Within the 10% precision of the FT-IR method, wheat germ 5S RNA exhibits essentially the same number of base pairs [14 A-U, 17 G-C, and 5 G-U; for a total of 36] in the absence of Mg2+ as in the presence of Mg2+ [14 A-U, 18 G-C, and 3 G-U; for a total of 35], in agreement with the UV hyperchromism estimate of G-C/(A-U + G-C) = 0.58.(ABSTRACT TRUNCATED AT 250 WORDS)     

New C-nucleoside analogs by dehydration of 1-benzyl-4,5,6,7-tetrahydro-6,6-dimethyl-2-(d-galacto-pentitol-1-yl)-indol-4-one

The reaction between 2-(benzylamino)-2-deoxy-d-glycero-l-gluco-heptose and 5,5-dimethyl-1,3-cyclohexanedione yields 1-benzyl-4,5,6,7-tetrahydro-6,6-dimethyl-2-(d-galacto-pentitol-1-yl)-indol-4-one (2). Acid-catalyzed, intramolecular dehydration of 2 under kinetically controlled conditions gives 1-benzyl-4,5,6,7-tetrahydro-2-α-d-lyxofuranosyl-6,6-dimethylindol-4-one; the anomeric configuration of this compound is only suggested. When the dehydration reaction is conducted under thermodynamically controlled conditions, it produces a 1:1 mixture of the α- and β-d-lyxopyranosyl compounds. The structures of the new compounds were elucidated by chemical and physical methods.     

Determination of base pairing in ribonucleic acid by Fourier-transform infrared spectrometry: yeast ribosomal 5S ribonucleic acid

Fourier-transform infrared (FT-IR) spectra of yeast ribosomal 5S RNA have been acquired at several temperatures between 30 and 90 degrees C. The difference spectrum between 90 (bases unstacked) and 30 degrees C (bases stacked) provides a measure of base stacking in the RNA. Calibration difference spectra corresponding to stacking of G-C or A-U pairs are obtained from "reference" FT-IR spectra of poly(rG) X poly(rC) minus 5'-GMP and 5'-CMP or poly(rA) X poly(rU) minus 5'-AMP and 5'-UMP. The best fit linear combination of the calibration G-C and A-U difference spectra to the 5S RNA (90-30 degrees C) difference spectrum leads to a total of 25 +/- 3 base pairs (17 G-C pairs + 8 A-U pairs) for the native yeast 5S RNA in the absence of Mg2+. In the presence of Mg2+, an additional six base pairs are detected by FT-IR (one G-C and five A-U). FT-IR melting curve midpoints show that A-U and G-C pairs melt together (65 and 63 degrees C) in the presence of Mg2+ but A-U pairs melt before G-C pairs (47 vs. 54 degrees C) in the absence of Mg2+.