Oxidation of n-alkanes: Growth of Pseudomonas putida

Summary The induction of alkane hydroxylase activity was investigated in two strains of Pseudomonas putida with a view to the production of primary alcohols. n-Nonanol production rates (16.0 mol/g dry wt/h) with an alcohol dehydrogenase negative mutant P. putida PpS173 were considerably lower than might be expected from the growth of a wild type on n-alkane. Production of cells by fed-batch culture on n-nonane, with a specific alkane hydroxylase activity of 3.9 mmol/g/h, was considered most suitable for isolation of the alkane hydroxylase.     

Localization of the Thy-1 antigen to the surfaces of rat retinal ganglion cells

Indirect immunofluorescence has been used to localize the Thy-1 antigen to ganglion cell axons, the ganglion cell layer and the inner plexiform layer in cryostat sections of adult and neonatal rat retina. In similar immunofluorescence experiments monoclonal antibodies raised against the 200,000 molecular weight neurofilament polypeptide bound only to ganglion cell axons and processes in the outer plexiform layer.Less than 1% of cells dissociated from 8 day postnatal rat retina had superficial Thy-1 antigen demonstrable by immunofluorescence; these cells were generally large and their size spectrum was similar to that of ganglion cells $\text{in}$$\text{situ}$. After culture for 1 day many of these Thy-1 positive cells had generated neurofilament antigen.We conclude that Thy-1 is found chiefly or exclusively on ganglion cells of eight day retina, and may be useful in the identification and isolation of these cells by immunoselection procedures.     

Isolation and characterization of two individual glycoprotein components from human milk-fat-globule membranes.

Two individual glycoprotein components from human milk-fat-globule membranes (MFGM) has been purified by selectively extracting the membrane glycoproteins followed by lectin affinity chromatography and gel filtration on Sephadex G-200 in the presence of protein-disaggregating agents. The purified glycoprotein components, termed 'epithelial-membrane glycoprotein' (EMGP-155 and EMGP-39) have estimated molecular weights of 155 000 and 39 000 respectively, and yield a single band under reducing conditions on sodium dodecyl sulphate/polyacrylamide gel. EMGP-155 and EMGP-39 contain 21.0% and 7.0% carbohydrate by weight, with fucose (13.5%, 12.4%), mannose (3.7%, 6.2%), galactose (28.5%, 22.6%), N-acetylglucosamine (17.8%, 7.4%) and sialic acid (36.4%, 51.4%) of the carbohydrate moiety respectively. For both the glycoprotein components, aspartic and glutamic acid and serine are the major amino acid residues.     

Mechanistic studies on C-19 demethylation in oestrogen biosynthesis

Mechanistic aspects of the biosynthesis of oestrogen have been studied with a microsomal preparation from full-term human placenta. The overall transformation, termed the aromatization process, involves three steps using O₂ and NADPH, in which the C-19 methyl group of an androgen is oxidised to formic acid with concomitant production of the aromatic ring of oestrogen: [Formula: see text] To study the mechanism of this process in terms of the involvement of the oxygen atoms, a number of labelled precursors were synthesized. Notable amongst these were 19-hydroxy-4-androstene-3,17-dione (II) and 19-oxo-4-androstene-3,17-dione (IV) in which the C-19 was labelled with ²H in addition to ¹⁸O. In order to follow the fate of the labelled atoms at C-19 of (II) and (IV) during the aromatization, the formic acid released from C-19 was benzylated and analysed by mass spectrometry. Experimental procedures were devised to minimize the exchange of oxygen atoms in substrates and product with oxygens of the medium. In the conversion of the 19-[¹⁸O] compounds of types (II) and (IV) into 3-hydroxy-1,3,5-(10)-oestratriene-17-one (V, oestrone), it was found that the formic acid from C-19 retained the original substrate oxygen. When the equivalent ¹⁶O substrates were aromatized under ¹⁸O₂, the formic acid from both substrates contained one atom of ¹⁸O. It is argued that in the conversion of the 19-hydroxy compound (II) into the 19-oxo compound (IV), the C-19 oxygen of the former remains intact and that one atom of oxygen from O₂ is incorporated into formic acid during the conversion of the 19-oxo compound (IV) into oestrogen. This conclusion was further substantiated by demonstrating that in the aromatization of 4-androstene-3,17-dione (I), both the oxygen atoms in the formic acid originated from molecular oxygen. 10β-Hydroxy-4-oestrene-3,17-dione formate, a possible intermediate in the aromatization, was synthesized and shown not to be converted into oestrogen. In the light of the cumulative evidence available to date, stereochemical aspects of the conversion of the 19-hydroxy compound (II) into the 19-oxo compound (IV), and mechanistic features of the C-10–C-19 bond cleavage step during the conversion of the 19-oxo compound (IV) into oestrogen are discussed.     

Inhibition of selected fungi by bacterial isolates fromTripsacum dactyloides L.

Summary In an incubation study addition of green manure caused a reduction in the ammonia volatilized initially from both sodic and reclaimed soils but extensive volatilization occurred from the sodic soil, amended with green manure, after the tenth day till the conclusion of the experiment after 9 weeks. Volatilization from the reclaimed soil was much less. There was a slight build up of organic carbon and ammoniacal nitrogen in both the soils though greater in the reclaimed soil. More of nitrate and nitrite accumulated in the sodic soil.     

Adrenergic regulation of pyruvate kinase and gluconeogenesis in hepatocytes from phosphorylase kinase-deficient (gsdgsd) rats

Hepatocytes were prepared from a strain of rats deficient in hepatic phosphorylase $\text{b}$ kinase and were used to assess the role of this enzyme in the adrenergic regulation of pyruvate kinase and gluconeogenesis. Epinephrine (10 μM) stimulated glucose output and gluconeogenesis from 1.8 mM lactate but did not significantly affect the concentration of hepatocyte glycogen. In addition epinephrine treatment led to an inhibition of pyruvate kinase. The stimulation of gluconeogenesis and the inhibition of pyruvate kinase by epinephrine were blocked by both α- and β-antagonists: similar effects with epinephrine were observed in cells from control animals. It is concluded that mechanisms for the adrenergic regulation of pyruvate kinase and gluconeogenesis are similar in hepatocytes from both phosphorylase kinase-deficient and normal rats.     

Differential biosynthesis of polyunsaturated fatty acids by Tetrahymena supplemented with ergosterol.

Tetrahymena grown with foreign sterols such as ergosterol incorporate them into cellular membranes at the expense of the native compound, tetrahymanol. It is shown that cells grown with ergosterol have a lessened capacity to produce the polyunsaturated linoleic and gamma-linolenic acids from [14C]oleic acid. However, the same cells have normal capacities to introduce double bonds at C-6 into linoleate, alpha-linolenate, or cis-vaccenate. Thus, a presumed 12-desaturase is inhibited in the presence of ergosterol, while desaturation at C-6 is unaffected.     

Relationship of posture and age to urinary protein excretion.

The influence of posture and age on urinary protein excretion was studied in 120 normal men volunteers. The supine excretion rate was less than 140 mug/min in all but two people (median value 38 mug/min) and tended to increase with age. The excretion rate decreased on quiet standing in 80% of people, which corresponded to a fall in creatinine-clearance. In the remaining 20% protein excretion increased on standing but generally remained within normal limits and was dissociated from changes in creatinine clearances. This increase was more prevalent in younger people and may represent a phenomenon analogous to orthostatic proteinuria, differing only quantitatively.     

Matrix Metalloproteinase 9 Exerts Antiviral Activity against Respiratory Syncytial Virus

Increased lung levels of matrix metalloproteinase 9 (MMP9) are frequently observed during respiratory syncytial virus (RSV) infection and elevated MMP9 concentrations are associated with severe disease. However little is known of the functional role of MMP9 during lung infection with RSV. To determine whether MMP9 exerted direct antiviral potential, active MMP9 was incubated with RSV, which showed that MMP9 directly prevented RSV infectivity to airway epithelial cells. Using knockout mice the effect of the loss of Mmp9 expression was examined during RSV infection to demonstrate MMP9’s role in viral clearance and disease progression. Seven days following RSV infection, Mmp9 ^-/- mice displayed substantial weight loss, increased RSV-induced airway hyperresponsiveness (AHR) and reduced clearance of RSV from the lungs compared to wild type mice. Although total bronchoalveolar lavage fluid (BALF) cell counts were similar in both groups, neutrophil recruitment to the lungs during RSV infection was significantly reduced in Mmp9 ^-/- mice. Reduced neutrophil recruitment coincided with diminished RANTES, IL-1β, SCF, G-CSF expression and p38 phosphorylation. Induction of p38 signaling was required for RANTES and G-CSF expression during RSV infection in airway epithelial cells. Therefore, MMP9 in RSV lung infection significantly enhances neutrophil recruitment, cytokine production and viral clearance while reducing AHR.