Motility as a morphogenic character in the genus Arthrobacter.

Motility in Arthrobacter atrocyaneus, A. citreus, and A. simplex was found to correlate with the morphogenic cycle of these organisms. The percentage of the A. atrocyaneus and A. simplex populations that were flagellated at a given time during the growth cycle differed significantly from that of the normorphogenic Pseudomonas aeruginosa population. Flagellation in A. atrocyaneus was shown to be dependent upon the morphogenic cycle rather than upon growth. The commitment to flagellar synthesis in A. atrocyaneus was found to occur only after induction to the rod morphology. Flagellar synthesis in A. atrocyaneus was shown to be restricted to only a small segment of the morphogenic cycle.     

Sphere-rod morphogenesis in Arthrobacter crystallopoietes. II. Peptides of the cell wall peptidoglycan

Cell walls of Arthrobacter crystallopoietes grown as spheres and as rods were solubilized by treatment with the B enzyme from Chalaropsis, an N-acetylmuramidase. The neutral glycopeptides were then isolated by chromatography on ECTEOLA cellulose. The glycopeptides, consisting of disaccharide-peptide units interlinked by peptide cross-bridges, were fractionated by gel filtration on Sephadex columns into oligomers of various sizes. The size distribution ranged from monomers with no cross-bridges to polymers with a high degree of polymerization, but did not differ significantly between cell walls from cells grown as spheres or rods. Some small differences in the distribution of C- and N-terminal amino acids were found. Analyses revealed that all the peptide bridges in the glycopeptide fractions from rod cell walls were formed by one l-alanine residue. In sphere cell walls, l-alanine was also found, but, in addition, higher oligomers of the glycopeptide contained glycine in their cross-bridges. These results were confirmed by determinations of C- and N-terminal amino acids released after lysostaphin and AL-1 enzyme digestions and by Edman degradations. Models representing the structures of the sphere and rod cell walls are presented. These structures indicate that the sphere cell wall is probably a more loosely knit macromolecule than is the rod cell wall.     

Induction and regulation of neuraminidase synthesis in Arthrobacter sialophilus.

A variety of N-acetylneuraminic acid (AcNeu) derivatives and analogs were examined as inducers of the extracellular neuraminidase of Arthrobacter sialophilus. Neuraminidase inductions were primarily studied with tryptone-yeast extract-grown cells after washing and resuspension in a defined replacement medium. The addition of readily metabolizable carbon sources to the latter, such as 0.1% casein hydrolysate, glutamate, or glucose, enhanced enzyme synthesis. Enzyme appearance occurred after a lag in the uptake of inducers, suggesting the participation of a co-inducible transport system. Neuraminidase formation during exponential growth in the presence of AcNeu ceased after depletion of this end product from the medium. It was found, besides AcNeu, that its methyl ester, 2-deoxy-2,3-dehydro-N-acetylneuraminic acid and 2-deoxy-2,3-dehydro-N-acetyl-neuraminic acid methyl ester are each active inducers, whereas beta-anomers of AcNeu-ketosides are not. These results, in comparison to known enzyme specificity, have revealed significant differences and parallels between the inductive and catalytic processes for neuraminidase. In particular, it would appear that the free carboxylate and oxygenation at C-2 of AcNeu, essential for enzyme catalysis with traditional AcNeu substrates, are not necessary for induction and, furthermore, that transition state analogs can specifically induce this enzyme. The failure to observe catabolite repression in this system is discussed in relation to the intermediary metabolism of the genus Arthrobacter.     

Facultative methylotroph belonging to the genus Arthrobacter]

A bacterial strain has been isolated and identified, on the basis of its morphological and physiologo-biochemical properties, as Arthrobacter globiformis. The bacterium is a facultative methylotroph and grows not only on media with various organic compounds but also in the presence of methylated amines as a sole source of carbon, nitrogen, and energy. Other C1-substrates were not utilized.     

Peptidoglycan compositions of a new strain of Arthrobacter crystallopietes during sphere-rod morphogenesis.

Arthrobacter crystallopoieties ATCC 15481 was used to isolate a new strain. designated Arthrobacter crystallopoieties EPSR-16, which had a mass doubling time in brain heart infusion broth and in glucose/salts/yeast extract medium of 30 min compared to 2.40 h for the parent strain in similar media. The growth rates for the new strain and for the parent were close to 12 h in glucose/salts medium. The new strain formed well-separated cocci and diplococci in glucose/salts medium, and upon nutrient shift-up all the cells in the population gradually changed into well-separated rods of regular shape. In the spherical state the cell wall peptidoglycan of the new strain contained lysine and no diaminopimelic acid. A gradual loss in lysine and a gain in diaminopimelic acid occurred during morphogenesis. Diaminopimelic acid became predominant in the cell wall during balanced growth in the rod state.     

Macromolecular synthesis and cell division during morphogenesis of Arthrobacter crystallopoietes

The sphere-rod-sphere morphology cycle of Arthrobacter crystallopoietes was accompanied by changes in the rate of growth and the rates of DNA, RNA and protein synthesis. The patterns of macromolecule synthesis resembled those found in other bacteria during a step-up followed by a step-down in growth rate. During the step-up in growth spherical cells grew into rods and macromolecules were synthesized in the absence of cell division. During stepdown, successive rounds of septation produced progressively smaller cells which did not separate and remained in chains. The morphology of the cells was dependent on the growth rate and could be altered by changing the dilution rate in a malate-limited chemostat. Gradual transitions in morphology and gradual increases in macromolecule content of the cells occurred as the growth rate was increased in the chemostat. Sphere to rod morphogenesis occurred when DNA synthesis was inhibited by treatment with mitomycin C or by thymine starvation. The DNA-deficient rods did not divide and eventually lysed. DNA, RNA and protein synthesis were continuously required for the reductive division of rods to spheres.Abbreviations MS mineral salts - GS mineral salts plus glucose - CA casamino acids - GSCA mineral salts plus glucose plus casamino acids - cAMP cyclic adenosine-3,5-monophosphate - RNA ribonucleic acid - DNA deoxyribonucleic acid     

An investigation of morphogenesis within the genus Trifolium

Morphogenic responses within the genus Trifolium were investigated by culturing various explants from seedlings of 72 species. Seedlings from 32 species produced callus alone, 40 produced adventitious shoots and/or roots, of which 25 species produced only shoots and 7 species formed only roots. Seedlings within each species also varied in their response to culture. The section of these seedlings most likely to produce adventitious shoots was the original shoot with the remnants of the surrounding hypocotyl and cotyledons, followed by the excised cotyledons themselves.Inter- and intra-varietal variation was observed in T. repens. Genotypes that produced adventitious buds were selected and crossed. An improvement in the proportion of the population capable of morphogenesis was observed in one cultivar.     

Control of morphogenesis in Arthrobacter crystallopoiets: effect of cyclic adenosine 3'',5''-monophosphate.

The intracellular levels of cyclic adenosine 3',5'-monophosphate (cyclic AMP) were measured at various intervals during growth and morphogenesis in Arthrobacter crystallopoietes. Cyclic AMP levels remained relatively constant throughout growth in spherical cells grown in glucose-based media. Immediately after inoculation of spheres from glucose- to succinate-containing media, a 30-fold increase in intracellular cyclic AMP was detected. This dramatic rise in cyclic AMP preceded the observed change in cellular morphology from spheres to rods. The cyclic AMP level in rod-shaped cells rapidly dropped to a relatively stable concentration during the exponential growth phase. At the onset of stationary phase and rod-to-sphere morphological transition, a second peak of cyclic AMP was observed. Neither of these two peaks was detectable in a morphogenetic mutant that grew only as spheres. The intracellular levels of cyclic AMP in this mutant remained constant throughout exponential growth and decreased slightly during stationary phase. Effects of exogenously added cyclic nucleotides and their derivatives to both parent and mutant cultures were investigated. The data presented indicate that dramatic changes in intracellular cyclic AMP levels occur just before the morphological transitions characteristic of the morphogenetic cycle in A. crystallopoietes. It is suggested that cyclic AMP is a contributing factor in the regulatory phenomenon associated with morphogenesis in this bacterium.     

Electron microscopic study of cell surface rings during cell division and morphogenesis of Arthrobacter crystallopoietes.

The whole cell ultrastructure during cell division and morphogenesis of Arthrobacter crystallopoietes was monitored using electron microscopic techniques. Glucose-grown spherical cells were inoculated into succinate-based medium. In this medium, the organism undergoes a morphogenetic cycle consisting of elongation of spheres to rods, exponential growth as rods, and fragmentation of rods to spherical cells. Raised bands or rings that encircled the cells were evident on the cell surface of both sphere- and rod-shaped cells. Many rod-shaped cells possessed two or more rings arranged adjacent to each other in a parallel orientation. At each cell division a new ring was formed on both siblings. However, as predicted by the proposed model of unidirectional cell growth and by maintaining a ring from the previous generation, unequal numbers of rings were observed on sibling cells. Only one ring was visible on most of the spherical inoculum cells, but in some cases a second ring perpendicular to the other ring was observed. Parallel rings were found on spherical cells resulting from fragmentation or reductive cell division of rods during the stationary growth phase. Thus, these spheres could be distinguished from inoculum spheres containing a single ring or perpendicular orientation of rings. The number of rings per cell and arrangement of rings on the cell surface of sibling cells after cell division, but before cell separation, are discussed with respect to cell age, cell division, and sphere-rod-sphere morphogenesis of A. crystallopoietes.     

Induction and Purification of Endo-beta-N-Acetylglucosaminidase from Arthrobacter protophormiae Grown in Ovalbumin

Arthrobacter protophormiae produced a high level of extracellular endo-beta-N-acetylglucosaminidase when cells were grown in a medium containing ovalbumin. The enzyme was induced by the glycopeptide fraction of ovalbumin prepared by pronase digestion. Production of the enzyme was also induced by glycoproteins such as yeast invertase and bovine ribonuclease B but not by monosaccharides such as mannose, N-acetylglucosamine, and galactose. The enzyme was purified to homogeneity as demonstrated by polyacrylamide gel electrophoresis and has an apparent molecular weight of about 80,000. The enzyme showed a broad optimum pH in the range of pH 5.0 to 11.0. The enzyme hydrolyzed all heterogeneous ovalbumin glycopeptides, although the hydrolysis rates for hybrid type glycopeptides were very low. The substrate specificity of A. protophormiae endo-beta-N-acetylglucosaminidase was very similar to that of Endo-C(II) from Clostridium perfringens. Therefore, the enzyme induction by A. protophormiae seems to have a close relation to the substrate specificity of the enzyme.     

Induction of d-6-hydroxynicotine oxidase in resting cells of Arthrobacter oxidans

Resting cell suspensions of Arthrobacter oxidans were shown to synthesize the inducible enantiozyme, d-6-hydroxynicotine oxidase, in the presence of d-nicotine or d-6-hydroxynicotine. The corresponding l-enantiomers, as well as -methylaminopropyl-(6-OH-pyridyl-3)-ketone, which is the product of the reaction catalyzed by the enzyme, were ineffective as inducers. l-6-Hydroxynicotine inhibited induction by d-nicotine and d-6-hydroxynicotine while l-nicotine inhibited induction by d-6-hydroxynicotine and had no effect on induction by d-nicotine. Enzyme induction was also found to be inhibited by glucose, 2-deoxy-d-glucose and by several intermediates of the tricarboxylic acid cycle. An absolute requirement for protein synthesis and for oxygen was also demonstrated to be necessary for the reactions involved in the covalent attachment of flavin adenine dinucleotide to pre-existing precursor protein to yield the catalytically active d-6-hydroxynicotine oxidase.H. C. R. completed these studies while on sabbatical leave from the Department of Botany and Microbiology, Arizona State University, Tempe, Arizona 85281, U.S.A.     

节杆菌M3黄嘌呤氧化酶的诱导效应研究

目的:寻找高效廉价的诱导物,并研究其诱导条件,以期进一步提高节杆菌M3(Anhrobacter M3)产黄嘌呤氧化酶(xan-thine oxidase,简写为XOD)水平.方法:以产酶水平和生产成本为标准,比较了几种化合物对Arthrobacter M3的诱导产酶的效果.选出最适诱导物,并对其诱导条件进行了研究.结果:几种化合物中,次黄嘌呤的诱导效果最佳.对次黄嘌呤诱导效应的研究发现,在发酵前期加入3g/L的次黄嘌呤对产黄嘌呤氧化酶的诱导效果最好.节杆菌M3降解次黄嘌呤的代谢产物中,尿素的存在对XOD的合成有强的抑制作用,而添加甘氨酸能促进XOD的产生.结论:在最佳条件下,次黄嘌呤诱导节杆菌M3产酶水平728.33U/L,超过了国内所报道的最高水平.     

Myceloid growth of Arthrobacter globiformis and other Arthrobacter species.

Transitory myceloid growth occurs in certain complex media with Arthrobacter globiformis strain ATCC 8010. This type of growth, however, was not observed in a medium which contained an array of metal ions but did not contain agents able to complex metal ions. Addition of metal-complexing agents to this medium caused an interruption in the life cycle of strain 8010 so that growth occurred only as the myceloid form. It appeared that manganese was the critical metal that was removed by the metal-complexing agents. During growth, the myceloid cells started to fragment, but wall septation was incomplete. A. globiformis strain ATCC 4336 and several other Arthrobacter species and soil isolates, but not Arthrobacter crystallopoietes, responded to metal-complexing agents as did strain 8010. Biotin and vitamin B12 were not involved in this myceloid growth.     

Induction of cellular morphogenesis in Myxococcus xanthus. II. Macromolecular synthesis and mechanism of inducer action

Sadler, William (University of Minnesota, Minneapolis), and Martin Dworkin. Induction of cellular morphogenesis in Myxococcus xanthus. II. Macromolecular synthesis and mechanism of inducer action. J. Bacteriol. 91:1520-1525. 1966.-Net changes in ribonucleic acid (RNA), deoxyribonucleic acid (DNA), and protein syntheses in cells of Myxococcus xanthus during induced, synchronous conversion to microcysts are described. The net synthesis of all three macromolecules was temporarily halted for a brief period during the initiation of shape change. Synthesis then resumed and leveled off when refractile microcysts began to appear. The conversion was completely sensitive, throughout the process, to low concentrations of chloramphenicol and actinomycin D. The uptake of amino acids and uracil was linear throughout the conversion, suggesting that the plateaus in rates of net synthesis of protein and RNA represented a period of rapid turnover. The most effective inducers of microcyst formation were fully saturated aliphatic compounds containing 2 to 4 carbon atoms and at least one primary or secondary alcohol group. Studies with labeled inducer indicated that the inducer need not be taken up by the cells to be effective, and probably interacts with some peripheral structure of the cell. The possibility that induction involves an alteration of a membrane-DNA complex is discussed.     

Alteration of glucose metabolism of Arthrobacter crystallopoietes by compounds which induce sphere to rod morphogenesis

Succinate and several other compounds which induce sphere to rod morphogenesis of A. crystallopoietes were found to suppress both catabolism and assimilation of glucose. Diauxic growth patterns resulted from growth on glucose plus any one of these compounds. Glutamate stimulated growth but was not an inducer of morphogenesis. With this compound, diauxic growth and suppression of glucose catabolism or assimilation did not occur. Glucose permease was studied with alpha-methylglucoside as substrate. The entry system for glucose was found to involve active transport and to have a K(m) of 8 x 10(-4)m. It was inducible, was repressed in succinate-grown cells, and was also inhibited by succinate. The exit system was constitutive and appeared to be less sensitive than the entry system to inhibition by azide. The properties of the glucose permease system may account for the slow growth of the organism on glucose and the preferred use of other substrates for growth. Studies of metabolic pathways for glucose metabolism indicated the operation of the Embden-Meyerhof-Parnas (EMP) and pentose phosphate pathways and of the tricarboxylic acid cycle. Cells grown on glucose plus limiting amounts of succinate or other inducers of morphogenesis metabolized the glucose only after exhaustion of the inducers. Under these circumstances, the organisms employed the EMP pathway to a greater extent than when growing on glucose as sole carbon source.