Cytokinin activity: localization in transfer RNA preparations

Transfer RNA from yeast, liver, and Escherichia coli has cytokinin activity in the tobacco callus bioassay, whereas ribosomal RNA from yeast is inactive. In contrast to fractions of yeast transfer RNA rich in serine acceptor and cytokinin activity, preparations (70 to 90 percent pure) of arginine transfer RNA(2), glycine transfer RNA, phenylalanine transfer RNA, and valine transfer RNA(1) and of highly purified alanine transfer RNA from yeast were inactive at concentrations of 20 to 2500 micrograms per liter. One molecule of 6-(gamma,gamma-dimethylallylamino) purine per 20 molecules of yeast tRNA would account for the observed cytokinin activity. The number of major molecular species contributing to cytokinin activity of transfer RNA, therefore, must be small.     

X-ray diffraction patterns of transfer RNA consistent with the presence of short parallel helices in the molecule

X-ray diffraction data of yeast formylmethionine transfer RNA, Escherichia coli phenylalanine transfer RNA, and Escherichia coli arginine transfer RNA single crystals are compared with the Fourier transform of a helix. The results are consistent with the presence of short parallel double helical segments in the transfer RNA molecules.     

Stathmin-tubulin interaction gradients in motile and mitotic cells

The spatial organization of the microtubule cytoskeleton is thought to be directed by steady-state activity gradients of diffusible regulatory molecules. We visualized such intracellular gradients by monitoring the interaction between tubulin and a regulator of microtubule dynamics, stathmin, using a fluorescence resonance energy transfer (FRET) biosensor. These gradients were observed both during interphase in motile membrane protrusions and during mitosis around chromosomes, which suggests that a similar mechanism may contribute to the creation of polarized microtubule structures. These interaction patterns are likely to reflect phosphorylation of stathmin in these areas.     

Identification of the cystic fibrosis gene: genetic analysis

Approximately 70 percent of the mutations in cystic fibrosis patients correspond to a specific deletion of three base pairs, which results in the loss of a phenylalanine residue at amino acid position 508 of the putative product of the cystic fibrosis gene. Extended haplotype data based on DNA markers closely linked to the putative disease gene locus suggest that the remainder of the cystic fibrosis mutant gene pool consists of multiple, different mutations. A small set of these latter mutant alleles (about 8 percent) may confer residual pancreatic exocrine function in a subgroup of patients who are pancreatic sufficient. The ability to detect mutations in the cystic fibrosis gene at the DNA level has important implications for genetic diagnosis.     

Angiotensin I: metabolism by plasma membrane of lung

(8-L-[(14)C] phenylalanine) angiotensin I is metabolized in one passage through blood-free lungs. Approximately 20 percent of the radioactivity emerges as angiotensin 11, the remainder as lower homologs. Radioactivity is not retained by the lungs but has the same volume of distribution and mean transit time as blue dextran, a compound unlikely to leave the intravascular space. Plasma membrane fractions of lung are capable of converting angiotensin I to angiotensin II. These data, taken together, indicate the circulating angiotensin I is metabolized by enzymes of the luminal surface of pulmonary endothelial cells.     

p-Chlorophenylalanine-induced chemical manifestations of phenylketonuria in rats

p-Chlorophenylalanine, a potent inhibitor of phenylalanine hydroxylase in vivo, has been used to simulate phenylketonuria in rats. This inhibitor, when administered with phenylalanine, produces marked elevation of blood and tissue phenylalanine without an increase in tyrosine. Disproportionate ratios of phenylalanine to tyrosine are characteristic of phenylketonuria in humans. The use of p-chlorophenylalanine permits the production of this characteristics amino acid imbalance in experimental animals.     

An ATP gate controls tubulin binding by the tethered head of kinesin-1

Kinesin-1 is a two-headed molecular motor that walks along microtubules, with each step gated by adenosine triphosphate (ATP) binding. Existing models for the gating mechanism propose a role for the microtubule lattice. We show that unpolymerized tubulin binds to kinesin-1, causing tubulin-activated release of adenosine diphosphate (ADP). With no added nucleotide, each kinesin-1 dimer binds one tubulin heterodimer. In adenylyl-imidodiphosphate (AMP-PNP), a nonhydrolyzable ATP analog, each kinesin-1 dimer binds two tubulin heterodimers. The data reveal an ATP gate that operates independently of the microtubule lattice, by ATP-dependent release of a steric or allosteric block on the tubulin binding site of the tethered kinesin-ADP head.     

The mitotic spindle: a self-made machine

The mitotic spindle is a highly dynamic molecular machine composed of tubulin, motors, and other molecules. It assembles around the chromosomes and distributes the duplicated genome to the daughter cells during mitosis. The biochemical and physical principles that govern the assembly of this machine are still unclear. However, accumulated discoveries indicate that chromosomes play a key role. Apparently, they generate a local cytoplasmic state that supports the nucleation and growth of microtubules. Then soluble and chromosome-associated molecular motors sort them into a bipolar array. The emerging picture is that spindle assembly is governed by a combination of modular principles and that their relative contribution may vary in different cell types and in various organisms.     

Selective inhibition of tubulin synthesis by amiprophos methyl during flagellar regeneration in Chlamydomonas reinhardi

Amiprophos methyl (APM) is a strong, readily reversible and highly selective inhibitor of tubulin synthesis in Chlamydomonas reinhardi. The extensive induction of tubulin synthesis that accompanies flagellar regeneration in this organism is prevented by 3 to 10 micrometerAPM. When applied after induction has begun, APM causes a rapid cessation of tubulin synthesis. Translation studies in vitro indicate that the lack of tubulin production in APM-treated cells is not due to a direct inhibition of tubulin messenger RNA translation but rather to a selective depletion of tubulin messenger RNA.     

Mapping of deletions and substitutions in heteroduplex DNA molecules of bacteriophage lambda by electron microscopy

Electron microscopy of heteroduplex DNA molecules, composed of one strand of Escherichia coli phage lambda(+) DNA annealed to the complementary DNA strand of a lambda deletion or substitution mutant, permits visualization, as well as precise measurements and mapping, of the unpaired single-stranded regions of nonhomology in the otherwise double-stranded molecules. In the lambdab2 mutant, the central segment (13 percent) of the lambda(+) DNA molecule is shown to be deleted. In the hybrid phages lambda(i434) and lambda(i21) a segment of the right arm of the lambda(+) genome (5.5 or 7.6 to 9 percent) is replaced by the corresponding immunity regions of phage 434 (3.3 percent or phage 21 (4 percent) DNA. The b5 region in the lambdab5 mutant appears to be identical to the i(21) segment. From these data it is possible to estimate the size and posiion of those lambda genes which are replaced by the i(434) and i(21) segments. The method permits preparing complete physical maps of viral genomes with a precision heretofore unattainable.     

Aromatic-aromatic interaction: a mechanism of protein structure stabilization

Analysis of neighboring aromatic groups in four biphenyl peptides or peptide analogs and 34 proteins reveals a specific aromatic-aromatic interaction. Aromatic pairs (less than 7 A between phenyl ring centroids) were analyzed for the frequency of pair type, their interaction geometry (separation and dihedral angle), their nonbonded interaction energy, the secondary structural locations of interacting residues, their environment, and their conservation in related molecules. The results indicate that on average about 60 percent of aromatic side chains in proteins are involved in aromatic pairs, 80 percent of which form networks of three or more interacting aromatic side chains. Phenyl ring centroids are separated by a preferential distance of between 4.5 and 7 A, and dihedral angles approaching 90 degrees are most common. Nonbonded potential energy calculations indicate that a typical aromatic-aromatic interaction has energy of between -1 and -2 kilocalories per mole. The free energy contribution of the interaction depends on the environment of the aromatic pair. Buried or partially buried pairs constitute 80 percent of the surveyed sample and contribute a free energy of between -0.6 and -1.3 kilocalories per mole to the stability of the protein's structure at physiologic temperature. Of the proteins surveyed, 80 percent of these energetically favorable interactions stabilize tertiary structure, and 20 percent stabilize quaternary structure. Conservation of the interaction in related molecules is particularly striking.     

Is there any chlorine monoxide in the stratosphere?

A ground-based search for stratospheric chlorine monoxide was carried out during May and October 1981 with an infrared heterodyne spectrometer in the solar absorption mode. Lines due to stratospheric nitric acid and tropospheric carbonyl sulfide were detected at about 0.2 percent absorptance levels, but the expected 0.1 percent lines of chlorine monoxide in this same region were not seen. Stratospheric chlorine monoxide is less abundant by at least a factor of 7 than is indicated by in situ measurements, and the upper limit for the integrated vertical column density of chlorine monoxide is 2.3 x 10(13) molecules per square centimeter at the 95 percent confidence level. These results imply that the release of chlorofluorocarbons may be significantly less important for the destruction of stratospheric ozone than is currently thought.     

Coordinated nonvectorial folding in a newly synthesized multidomain protein

The low-density lipoprotein receptor (LDL-R) is a typical example of a multidomain protein, for which in vivo folding is assumed to occur vectorially from the amino terminus to the carboxyl terminus. Using a pulse-chase approach in intact cells, we found instead that newly synthesized LDL-R molecules folded by way of "collapsed" intermediates that contained non-native disulfide bonds between distant cysteines. The most amino-terminal domain acquired its native conformation late in folding instead of during synthesis. Thus, productive LDL-R folding in a cell is not vectorial but is mostly posttranslational, and involves transient long-range non-native disulfide bonds that are isomerized into native short-range cysteine pairs.     

Structure of langmuir-blodgett films of disk-shaped molecules determined by atomic force microscopy

Monolayer Langmuir-Blodgett films of a discotic mesogen have been studied with atomic force microscopy (AFM). These measurements confirm the "edge on" arrangement for the disk-shaped molecules suggested by surface pressure-area isotherms and show that the molecules form columns that are separated by 17.7 angstroms +/- 10 percent. Column alignment is found to be predominantly along the film deposition direction, with an angular spread of 35 degrees . The AFM images also show that the mean disk separation within the columns is 5.1 +/- 1.3 angstroms, in good agreement with x-ray diffraction (XRD) results. Roomtemperature XRD measurements on bulk samples of the same material indicate a disordered-hexagonal liquid crystalline mesophase, with a column-to-column spacing of 19.9 +/- 0.2 angstroms.     

Stabilizing isopeptide bonds revealed in gram-positive bacterial pilus structure

Many bacterial pathogens have long, slender pili through which they adhere to host cells. The crystal structure of the major pilin subunit from the Gram-positive human pathogen Streptococcus pyogenes at 2.2 angstroms resolution reveals an extended structure comprising two all-beta domains. The molecules associate in columns through the crystal, with each carboxyl terminus adjacent to a conserved lysine of the next molecule. This lysine forms the isopeptide bonds that link the subunits in native pili, validating the relevance of the crystal assembly. Each subunit contains two lysine-asparagine isopeptide bonds generated by an intramolecular reaction, and we find evidence for similar isopeptide bonds in other cell surface proteins of Gram-positive bacteria. The present structure explains the strength and stability of such Gram-positive pili and could facilitate vaccine development.     

烤烟上部6片叶一次采收对顶部3片叶烘烤质量的影响

研究了不同采收方式对烤烟顶部3片叶物理性状、化学成分和中性香气物质含量的影响.结果表明:与对照相比,处理Ⅱ(顶部3片叶八、九成黄时,上部6片叶一次采收,取顶部3片有效叶进行烘烤)叶片填充值、平衡含水率、叶质量和含梗率增加,厚度减小;处理Ⅱ有助于提高顶部3片烟叶水溶性总糖含量,降低淀粉、总氮和蛋白质含量;处理Ⅱ明显提高了顶部3片烟叶中性挥发性香气物质总量,其中类胡萝卜素降解产物、苯丙氨酸代谢产物、西柏烷类降解产物和新植二烯等各类致香物质含量均有不同程度的提高.     

Amino acid changes provoked by streptomycin in a polypeptide synthesized in vitro

The misincorporation of isoleucine provoked by streptomycin in a polyuridylate-directed incorporating system from Escherichia coli has been examined by a double-labeling technique with phenylalanine-(3)H and isoleucine-(14)C. The polypeptides synthesized are associated with 70S ribosomes and contain only phenylalanine in the absence of streptomycin, and both phenylalanine and isoleucine in the presence of the antibiotic. Results of acid hydrolysis and subsequent chymotryptic digestion of the polypeptides indicate that the misincorporated amino acid is inserted by peptide bonding with phenylalanine and is randomly distributed along the chain.