Peptidoglycan of Pseudomonas aeruginosa

A practical method for preparing peptidoglycan from Ps. aeruginosa and E. coli was devised. After bacterial cells were dissolved in boiling 4% SDS solution, peptidoglycan was collected and washed with water by centrifugation. Peptidoglycan was treated further with pronase and lyophilized. The final preparation of peptidoglycan from Ps. aeruginosa appeared as a filmy coagulation in electron micrograph and its amino acid composition was determined as follows: Glu/Ala/A₂pm/Mur/GlcN (100/183/104/61/98). The lysozyme digest showed the same pattern as that of E. coli peptidoglycan. N-Terminal analysis suggested that about half of the peptide chains was interbridged by the peptide bond between Ala and A₂pm. The probable ratio of muropeptides in the peptidoglycan was estimated.     

Development of a Host-vector System for Gluconobacter suboxydans

A shuttle vector for Gluconobacter suboxydans and Escherichia coli was constructed by ligation of a cryptic plasmid, pMV201, found in G. suboxydans IFO 3130 to E. coli plasmid pACYC177. The chimeric plasmid named pMGlOl carries the ampicillin resistance gene derived from pACYC177 and transforms G. suboxydans var. α IFO 3254 as well as E. coli. The transformation conditions for G. suboxydansvar. α IFO 3254 were examined using pMGlOl DNA. Competent cells were induced efficiently by treatment with LiCl or RbCl CaCl₂ which induced the competency of Acetobacter was much less effective. Addition of polyethylene glycol enhanced the transformation efficiency significantly. An efficiency of approximately 10² transformants per μg DNA was finally obtained.     

Effect of Various Inhibitors on Lipase Action of Thermophilic Fungus Humicola lanuginosa S-38

The hydrolysis of olive oil by the Humicola lipase was inhibited by the addition of n-alcohols, fatty acids and surface active agents. The inhibition of n-alcohols was overcomed by the addition of more substrate but not by the addition of more enzyme. The inhibition of fatty acids and bile salts was eliminated by adding calcium ion. It was concluded that the inhibition of the Humicola lipase by n-alcohols, fatty acids and bile salts was not due to inactivation of the enzyme directly but due to the displacing of the substrate from the oil/water interface, thus blocking the enzyme from the substrate.     

A Lignan-type Stress Compound in Potato Infected with Nematode (Globodera rostochiensis)

Rustmicin, a new antibiotic active against the wheat stem rust fungus, was isolated from a cultured broth of Micromonospora chalcea 980-MC₁. Rustmicin showed strong inhibitory activity against the wheat stem rust fungus both in vitro and in pot tests in a greenhouse with MIC being 1 and 0.8/ig/ml, respectively. Its structure was determined by NMR spectroscopy to be a new 14-membered macrolide antibiotic lacking sugar substituents.     

Possible Mechanism of Oxygen Activation in Linoleate Peroxidation by Fusarium Lipoxygenase

The Oxygen activating mechanism of Fusarium lipoxygenase, a heme-containing dioxygenase, was studied. The enzyme did not require any cofactors, such as H₂O₂, however, both superoxide dismutase and catalase inhibited linoleate peroxidation by Fusarium lipoxygenase. A low concentration of H₂O₂ caused a distinct acceleration in enzymatic peroxidation. These results indicate that both O₂? and H₂O₂ are produced as essential intermediates of oxygen activation during formation of linoleate hydroperoxides by Fusarium lipoxygenase. This peroxidation reaction was also prevented by scavengers of singlet oxygen (¹O₂), but not by scavengers of hydroxy 1 radical (OH). Generation of O₂? in the enzyme reaction was detected by its ability to oxidize epinephrine to adrenochrome. Moreover, the rate of peroxide formation was greater in the D₂O than in the H₂O buffer system. These results suggest that the Haber–Weiss reaction (O₂^?+H₂O₂→OH^?+OH^·+¹O₂) is taking part in linoleate peroxidation by Fusarium lipoxygenase, and the ¹O₂ evolved could be responsible for the peroxidation of linoleate. H₂O₂ produced endogenously in the enzyme reaction might act as an activating factor for the enzyme. This possible mechanism of oxygen activation can explain the absence of a need for exogenous cofactors with Fusarium lipoxygenase in contrast to an other heme-containing dioxygenase, tryptophan pyrrolase, which requires an exogenous activating factor, such as H₂O₂.     

2-Nitropropane Dioxygenase from Hansenula mrakii: Re-characterization of the Enzyme and Oxidation of Anionic Nitroalkanes

2-Nitropropane dioxygenase, purified to homogeneity by an improved method from a yeast, Hansenula mrakii, has a molecular weight of 42,000, and consists of a single polypeptide. The enzyme contains 1 mol of FAD per mol of enzyme. The iron protein associated with previous preparations was removed by the present purification procedures. The enzyme catalyzes the oxygenative denitrification of anionic nitroalkanes much more effectively than that of the neutral ones with the optimum pH of 6.5. The Michaelis constants for the anionic substrates are as follows: 2-nitropropane, 1.61 mM; 1-nitropropane, 3.23 mM; nitroethane, 3.13 mM, and 3-nitro-2-butanol, 0.59 mM.     

Identification and Growth Characteristics of α-Galactosidase-producing Microorganisms

Two kinds of α-galactosidase-producing microorganisms, strain No. 31–2 and strain No. 7–5, have been isolated from soil and subjected to a determinative study. On the basis of the morphological and physiological characters, the strain No. 31–2 was identified to be belonged to genus Micrococcus and the strain No. 7–5 to genus Bacillus. The former strain, Micrococcus sp. No. 31–2, produced exclusively an intracellular α-galactosidase, and the latter one, Bacillus sp. No. 7–5, secreted the enzyme into culture medium. The cell growth and enzyme production of both strains were observed to reach the maximum under an alkaline culture condition. The intracellular α-galactosidase of Micrococcus sp. No. 31–2 was inducible by galactose, melibiose, and raffinose, while the α-galactosidase of Bacillus sp. No. 7–5 was produced constitutively.     

Effect of Exogenous Fatty Acids on the Cellular Fatty Acid Composition and Neomycin Formation in a Mutant Strain of Streptomyces fradiae

A mutant of Streptomyces fradiae which requires oleic acid for neomycin formation was isolated and the effects of exogenous fatty acids and other additives on the formation of neomycin were studied. Palmitic acid and high concentration of sodium ions could replace oleic acid in neomycin formation. The fatty acid spectrum of the mutant strain ST–5B was quite different from that of the parent strain 3123. The major fatty acid components of the mutant and the parent were anteiso 15:0 and iso 16: 0, respectively. However the fatty acid composition of the mutant was changed from the anteiso 15: 0-type to the parental iso 16: 0-type by the supplement of oleic acid or high concentration of sodium ions in the medium. In the case of palmitic acid, the major fatty acid component of the mutant cells was changed from anteriso 15: 0 to normal 16:0. The role of these additives in neomycin formation by the mutant is discussed.