Cloning and sequencing of the peroxisomal amine oxidase gene from Hansenula polymorpha

We have cloned the AMO gene, encoding the microbody matrix enzyme amine oxidase (EC 1.4.3.6) from the yeast Hansenula polymorpha. The gene was isolated by differential screening of a cDNA library, immunoselection, and subsequent screening of a H. polymorpha genomic library. The nucleotide sequence of a 3.6 kilobase stretch of DNA containing the amine oxidase (AMO) gene was determined. The AMO gene contains an open reading frame of 692 amino acids, with a relative molecular mass of 77,435. The 5' and 3' ends of the gene were mapped and show that the transcribed region measures 2134 nucleotides. The derived amino-acid sequence was confirmed by sequencing an internal proteolytic fragment of the purified protein. Amine oxidase contains the tripeptide sequence Ser-Arg-Leu, located 9 residues from the carboxy terminus, which may represent the topogenic signal for protein import into microbodies.     

Studies on modulation of the effects of colchicine by L-cysteine using bone marrow of Swiss mice

Colchicine (COL) elevates the frequency of micronucleated polychromatic erythrocytes (PE), the ratio of normochromatic to polychromatic erythrocytes (N/PE) and the frequency of large PE due to spindle disruption. Simultaneous i.p. injection of L-cysteine (CYS) does not influence the effects of COL while if administered 1 h prior to COL, CYS suppresses the N/PE ratio and frequency of large PE but not the frequency of micronucleated PE elevated by COL. Preincubation of CYS with COL at 37 degrees C for 1 h results in a significant decrease in all the COL effects. The modulatory effect of exogenous CYS appears to be due to its competition with the endogenous tubulin cysteine residues for interacting with COL.     

Effect of dibutyryl cyclic AMP on lipid synthesis in Microsporum gypseum

The influence of intracellular levels of cAMP on the lipid synthesis of Microsporum gypseum has been examined by exogenous supplementation of dibutyryl cAMP and its activators/inhibitors. Incorporation of [14C]acetate into various lipid fractions of M. gypseum was markedly enhanced in the presence of dibutyryl cAMP and its modulators, probably as a consequence of increased intracellular cAMP levels, which, in turn, affected the lipid biosynthesis. Increased activities of phosphatidic acid phosphatase, glycerol kinase, ethanolamine kinase and choline kinase in the presence of these additives supports the enhanced synthesis of phospholipids and suggests that lipid biosynthesis is being controlled by cAMP in M. gypseum.     

Alkaloid production byClaviceps sp. SD-58; Involvement of phosphatase isozymes

Simultaneous reduction in alkaloid yield and level of phosphatases by high concentrations of phosphate was observed inClaviceps sp. SD-58. Tryptophan-induced culture showed an increase in alkaloid yield and the level of phosphatases. Phosphate caused repression of both acid phosphatase (isoenzyme I) and alkaline phosphatase (isoenzymes III and V).     

Surfactant-induced breakthrough effects during the operation of two-phase biocatalytic membrane reactors

Surface-active components, both reactants and products, are frequently encountered in two-phase, aqueous-organic, biocatalytic reactions, When such reaction are carried out in a membrane reactor, employing a membrane selectively wetted by one of the two reactants, changes in the content of these surfactants- as a consequence of the progress of the reaction-can lead to wetting transitions at the two membrane-liquid interfaces as a result of adsorption of the tenside. This can lead to a decrease in the pressure required to cause the, initially, nonwetting phase to break through the membrane. Such effects render difficult the operation of two-phase membrane bioreactors. Hence, it is necessary to make a careful selection of the membrane material and type by considering factors such as UF versus MF and low MWCO versus high MWCO to enable the reactor to be operated without breakthrough, but without significantly compromising the reaction rates that can be maintained.The phenomena leading to breakthrough effects are discussed in this paper, and experimental results for the hydrolysis of ethyl laurate by lipase from Candida rugosa in a batch flat sheet membrane reactor are presented with the reactor operated with a variety of membranes. An experimental result showing the decrease in the pressure required to cause breakthrough of the organic phase (for the system ethyl laurate-lauric acid-water) as the content of the highly surface-active lauric acid in the organic phase is increased is also presented for an asymmetric, hydrophilic meta-aramid ultrafiltration membrane. (c) 1994 John Wiley & Sons, Inc.     

Detection of vitamin B12 and pantothenic acid in cell exudates of blue-green algae

The blue-green algaeChroococcus, Oscillatoria and Nostoc, excrete vitamin B₁₂ and pantothenic acid in the culture filtrates during the phase of active growth. The excretion has an ecological significance.     

Micro determination of polyamines with snake venom oxidase.

An accurate micromethod suitable for the assay of polyamines in concentrations of 2 to 30 nmol is described. It is based on the oxidation of polyamines by purified fractions of crude l-amino acid oxidase from Russell's viper venom. By a combination of two enzyme fractions, one which oxidizes polyamines and amino acids (AAP) and another which oxidizes only amino acids (AA), the technique can accurately determine polyamine concentrations in extracts of sera which may not be free of amino acids. Experiments described show 90 to 100% recovery of added polyamines in the presence of varying amounts of amino acids. Polyamines added to serum also showed recoveries ranging from 96 to 98%. The enzymes do not oxidize histamine and epinephrine and are very stable. The method does not require sophisticated equipment and is suitable for screening of large number of clinical samples to assess the importance of polyamines as a diagnostic test or their prognostic value in diseases like cancer.