Superficial Macromolecular Arrays on the Cell Wall of Spirillum putridiconchylium

Electron microscopy of the cell envelope of Spirillum putridiconchylium, using negatively stained, thin-sectioned, and replicated freeze-etched preparations, showed two superficial wall layers forming a complex macromolecular pattern on the external surface. The outer structured layer was a linear array of particles overlying an inner tetragonal array of larger subunits. They were associated in a very regular fashion, and the complex was bonded to the outer, pitted surface of the lipopolysaccharide tripartite layer of the cell wall. The relationship of the components of the two structured layers was resolved with the aid of optical diffraction, combined with image filtering and reconstruction and linear and rotary integration techniques. The outer structural layer consisted of spherical 1.5-nm units set in double lines determined by the size and arrangement of 6- by 3-nm inner structural layer subunits, which bore one outer structural layer unit on each outer corner. The total effect of this arrangement was a double-ridged linear structure that was evident in surface replicas and negatively stained fragments of the whole wall. The packing of these units was not square but skewed by 2 degrees off the perpendicular so that the "unit array" described by optical diffraction and linear integration appeared to be a deformed tetragon. The verity of the model was checked by using a photographically reduced image to produce an optical diffraction pattern for comparison with that of the actual layers. The correspondence was nearly perfect.     

Isolation and Partial Characterization of Escherichia coli Mutants with Low Levels of Transfer Ribonucleic Acid Nucleotidyltransferase

To determine the function of the enzyme transfer ribonucleic acid (tRNA) nucleotidyltransferase in vivo, five mutants of Escherichia coli containing low levels of this enzyme were isolated. Since no selection procedure for such mutants existed, these strains were isolated by assay of large numbers of colonies from a heavily mutagenized stock. A procedure employing cells made permeable to tRNA and ATP was used to screen the large number of colonies required for the isolation. All the mutants contained less than 20% of the normal level of the AMP-incorporating activity of tRNA nucleotidyltransferase in extracts prepared by several methods, and the best mutant contained only about 2% of this activity. Three of the mutants also had equally low levels of the cytidine 5'-monophosphate-incorporating activity of the enzyme. Despite these low activities, the mutant strains displayed relatively normal growth characteristics at all temperatures examined. The enzyme in the mutant strains was not temperature sensitive, nor were any other abnormal biochemical properties detected. tRNA isolated from the mutant strains was missing significant amounts of its 3' terminal adenosine 5'-monophosphate residue, amounting to 10 to 15% in the best mutant. However, only small amounts of the terminal cytidine 5'-monophosphate residue were missing. The results indicate that tRNA nucleotidyltransferase is involved in some aspect of synthesis or repair of the 3' terminus of tRNA, and that the enzyme is present in large excess over its requirements for this function.     

Preparation of Cells Permeable to Macromolecules by Treatment with Toluene: Studies of Transfer Ribonucleic Acid Nucleotidyltransferase

Transfer ribonucleic acid (tRNA) nucleotidyltransferase was studied after making cells permeable to macromolecules by treatment with toluene. The conditions of toluene treatment necessary for obtaining maximal activity were defined. Toluene treatment was most efficient when carried out for 5 min at 37 C at pH 9.0 on log-phase cells. No activity could be detected if cells were treated at 0 C, or in the presence of MgCl₂, or if the cells were in the stationary phase of growth. However, inclusion of lysozyme and ethylenediaminetetraacetic acid during the toluene treatment did render stationary phase cells permeable. The properties of tRNA nucleotidyltransferase from toluene-treated cells were essentially identical to those of purified enzyme with regard to pH optimum, specificity for nucleoside triphosphates and tRNA, and apparent K_m values for substrates. In addition to tRNA nucleotidyltransferase, a variety of other enzymes which incorporate adenosine 5′-triphosphate into acid-precipitable material could also be detected in toluene-treated cells. Centrifugation of cells treated with toluene revealed that tRNA nucleotidyltransferase leaked out of cells, whereas other activities remained associated with the cell pellets. Chromatography of the material extracted from toluene-treated cells on Sephadex G-100 indicated that toluene treatment selectively extracts lower molecular weight proteins. The usefulness of such a procedure as an initial step in purification of such enzymes, and its application to tRNA nucleotidyltransferase, is discussed.     

Conformational aspect of polypeptide structure. XLIV. Conformational transitions of poly(N-methyl-alanines) induced by trifluoroacetic acid

The synthesis of poly(N-methyl-L -alanine) and poly (N-methyl-DL -alanine) are described. The polymers were examined by 220 MHz high-resolution nuclear magnetic resonance (nmr) and circular dichroism (CD). The results demonstrate that poly(N-methyl-L -alanine) exists as an ordered helical structure with all the amide bonds in the trans configuration in appropriate solvents. As trifluoroacetic acid (TFA) is added to the solutions of the polymer in helix-supporting solvents, resonances corresponding to both trans and cis amide conformations of N-methyl, C-methyl, and α-CH are observed. The presence of both the trans and the cis peptide bonds in a polymer chain disrupts the ordered structures. Our conclusions from CD data are in agreement with the nmr results. Ultracentrifugation shows that degradation of the polymer chain does not occur during the TFA treatment.     

A re-evaluation of polypodium bradeorum and P. Colysoides

During field studies on the pteridophytes of Costa Rica, a peculiarly dimorphic polypodioid fern was found in the rain-forests of the Atlantic lowlands near Puerto Viejo. The variation in the fertile frond, ranging from simple and short petiolate to pinnatisect and long petiolate, coupled with peculiarly elongate and irregular sori, prompted further investigations. Additional herbarium specimens from localities in Mexico, British Honduras, Nicaragua, and Costa Rica showed intermixed variations between plants with all leaves simple (typified byPolypodium bradeorum Rosenstock) to plants with all leaves lobed or pinnatisect (typified byP. colysoides Maxon & Copeland). Other characters were judged sufficiently homogeneous to consider these individuals as conspecific underP. bradeorum. Morphological studies indicate parallel evolution of several characters in the Asiatic generaColysis, Microsorium, andLeptochilus on the one hand and the New World members ofMicro gramma, Pleopeltis, andPolypodium bradeorum on the other     

Taxonomy of carex sect. atratae (cyperaceae) in the southern rocky mountains

The sectionAtratae is highly differentiated in the southern Rocky Mountain region where the group has developed several endemic species. Eleven species are recognized, and three new combinations are proposed:C. parryana ssp.hallii, C. parryana ssp.idahoa, andC. norvegica ssp.stevenii.     

Chain conformation of copoly(γ-methyl D,L-glutamate)

Copolymers of γ-methyl D - and L -glutamates with various D /L ratios were prepared. Infrared absorption spectra of solid films were measured and sums of right- and left-handed helix contents were determined from intensities of amide V bands. Farultraviolet absorption spectra and optical rotatory dispersion of these copolymers in solutions are used to ascertain their helical character. Chain conformations of DL -copolypeptides are discussed.     

Purine metabolism in germinating wheat embryos

1. Both the acid-soluble fraction and the nucleic acid fraction of wheat embryos were extensively labelled after incubation for 6hr. in the presence of [8-(14)C]adenine. Subsequent incubation in the absence of labelled adenine resulted in no loss of radioactivity to the medium during a 48hr. period. Radioautography indicated that during this period there was a continuous increase in the radioactivity present in the acid-insoluble fractions of the root and leaf tissues relative to that present in the coleorhiza and coleoptile. 2. During incubation at 25 degrees there was a 26-fold increase in the activity of 3'-nucleotidase between 4hr. and 24hr.; the activities of enzymes hydrolysing AMP and IMP increased to a smaller extent. The activities of adenine phosphoribosyltransferase and hypoxanthine phosphoribosyltransferase increased three- to five-fold during incubation at 25 degrees for 24hr. 3. Adenosine kinase, inosine phosphorylase and 5-phosphoribosyl pyrophosphate synthetase activities were high in extracts from dry embryos and did not increase during 48hr. at 25 degrees . 4. The increase in 3'-nucleotidase activity was prevented by cycloheximide, cryptopleurine or incubation at 4 degrees , but not by actinomycin D; these treatments did not depress the activity of the other enzymes measured. 5. The results are discussed in relation to RNA translocation within the wheat embryo during germination.