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Biomedical subjects

J A Pateman

Publications and source records attributed to J A Pateman.

At least 19 recordsLinked to original sources
PubMed1985

Purification and preliminary characterization of alcohol dehydrogenase from Aspergillus nidulans.

Aspergillus alcohol dehydrogenase is produced in response to growth in the presence of a wide variety of inducers, of which the most effective are short-chain alcohols and ketones, e.g. butan-2-one and propan-2-ol. The enzyme can be readily extracted from fresh or freeze-dried cells and purified to homogeneity on Blue Sepharose in a single step by using specific elution with NAD+ and pyrazole. The pure enzyme has Mr 290 000 by electrophoresis or gel filtration; it is a homopolymer with subunit Mr 37 500 by electrophoresis in sodium dodecyl sulphate; its amino acid composition corresponds to Mr 37 900, and the native enzyme contains one zinc atom per subunit. The enzyme is NAD-specific and has a wide substrate activity in the forward and reverse reactions; its activity profile is not identical with those of other alcohol dehydrogenases.

Alcohol Dehydrogenase↗
PubMed1985

Genomic clones of Aspergillus nidulans containing alcA, the structural gene for alcohol dehydrogenase and alcR, a regulatory gene for ethanol metabolism.

Our aim was to obtain from Aspergillus nidulans a genomic bank and then clone a region we expected from earlier genetic mapping to contain two closely linked genes, alcA, the structural gene for alcohol dehydrogenase (ADH) and alcR, a positive trans-acting regulatory gene for ethanol metabolism. The expression of alcA is repressed by carbon catabolites. A genomic restriction fragment characteristic of the alcA-alcR region was identified, cloned in pBR322, and used to select from a genomic bank in lambda EMBL3A three overlapping clones covering 24 kb of DNA. Southern genomic analysis of wild-type, alcA and alcR mutants showed that the mutants contained extra DNA at sites near the center of the cloned DNA and are close together, as expected for alcA and alcR. Transcription from the cloned DNA and hybridization with a clone carrying the Saccharomyces cerevisiae gene for ADHI (ADC1) are both confined to the alcA-alcR region. At least one of several species of mature mRNA is about 1 kb, the size required to code for ADH. For all species, carbon catabolite repression overrides control by induction. The overall characteristics of transcription, hybridization to ADC1 and earlier work suggest that alcA consists of a number of exons and/or that the alcA-alcR region represents a cluster of alcA-related genes or sequences.

Alcohol Dehydrogenase↗
PubMed1983

A polyamine-sensitive mutant of Aspergillus nidulans.

A mutation designated spsA1 has been induced in the putrescine (puA2) auxotroph of Aspergillus nidulans which enables this mutant to grow on low concentrations of spermidine in place of putrescine. In addition, the spsA1 mutant, irrespective of putrescine requirement, is abnormally sensitive to high concentrations of spermidine, spermine or the polyamine analogue methylglyoxal bis(guanylhydrazone). When spsA1 strains are grown on medium containing spermidine, uptake of the polyamine continues at a high level for a longer period than in the wild-type and leads to a doubled intracellular spermidine pool. A similar increase in the intracellular spermine pool results from growth on spermine.

Aspergillus nidulans↗
PubMed1982

The regulation of urease activity in Aspergillus nidulans.

Aspergillus nidulans can utilize urea as a sole source of nitrogen but not as a carbon source. Urea is degraded by a urease. Mutation at any one of three genes, ureB, ureC, and ureD, may result in deficient urease activity. The ureB gene is closely linked to ureA, the structural gene for the urea transport protein. The heat lability of ureB- revertant strain, intragenic complementation tests, and the linkage of ureB to ureA suggest that ureB is the urease structural gene. The ureD gene is probably involved in the synthesis or incorporation of a nickel cofactor essential for urease activity. The function of the ureC gene is not known. Urease is not induced but is subject to nitrogen regulation. The urease activities of ammonium-derepressed mutants show that the effector of nitrogen regulation is more likely to be glutamine than ammonium. When glutamine is present in the medium, urease appears to be inactivated by some means which does not involve a newly synthesized protease or a direct interaction between glutamine and urease.

Aspergillus nidulans↗
PubMed1982

Urea and thiourea transport in Aspergillus nidulans.

Wild-type Aspergillus nidulans has an active transport system specific for urea which concentrates urea at least 50-fold relative to the extracellular concentration. It is substrate concentration dependent, with an apparent Km of 3 x 10-(5) M for urea. Competition studies and the properties of mutants indicate that thiourea is taken up by the same system as urea. Thiourea is toxic at 5mM to wild-type cells of Aspergillus nidulans. Mutants, designated ureA1 to ureA16, resistant to thiourea have been isolated, and transport assays and growth tests show that they are specifically impaired in urea transport. The mutant ureA1 has a higher Km value than the wild type for thiourea uptake. The ureA locus has been assigned to linkage group VIII. ureA1 is recessive for thiourea resistance while semidominant for the low uptake characteristic. The urea uptake system is under nitrogen regulation, with L-glutamine as the probable effector. The mutants, meaA8 and gdhA1, which are insensitive to ammonium control of many nitrogen-regulated metabolic systems, are also insensitive to ammonium control of urea uptake, but both are sensitive to L-glutamine regulation.

Aspergillus nidulans↗
PubMed1982

Polyamine transport in Aspergillus nidulans.

The uptake of putrescine, spermidine and spermine was studied in Aspergillus nidulans using 14C-labelled polyamines. Active transport systems, inhibited by azide and regulated by nitrogen availability, exist at least for putrescine and spermidine. Putrescine is taken up two to three times more rapidly than spermidine, reflecting a lower Km for the former substrate. The two uptake systems appear to be independent, spermidine uptake being inhibited by both putrescine and spermine, while putrescine uptake is not inhibited by the other two polyamines. The relationships of these transport systems to the tenfold or greater difference between spermidine and putrescine concentrations required to support growth of the putrescine auxotroph are discussed.

Aspergillus nidulans↗
PubMed1980

Nickel requirement of a urease-deficient mutant in Aspergillus nidulans.

The addition of nickel ions restored urease activity in vivo and ability to grow on urea in a mutant strain of Aspergillus nidulans otherwise unable to utilize urea. This train carries a mutation in the ureD locus, one of four loci involved in urea utilization. No other urease-deficient strains tested responded to the presence of nickel ions. The analogous characteristics of the ureD mutant and the nitrate reductase and xanthine dehydrogenase associated cnxE mutants in Aspergillus nidulans are discussed. It is postulated that the ureD locus is in some way involved in the production or incorporation of a nickel cofactor essential for urease activity.

Aspergillus nidulans↗
PubMed1979

The regulation of hexokinase and phosphoglucomutase activity in Aspergillus nidulans.

The levels of glucose-6-phosphate and 6-phosphogluconate dehydrogenase in wildtype cells of Aspergillus nidulans varied with the carbon and nitrogen source. In general, hexokinase activity did not vary with carbon or nitrogen source. The ammonium derepressed mutant amrA1 had only 50% of the wildtype level of hexokinase. Phosphoglucomutase activity was low in wildtype cells grown with nitrate, but high in cells grown with ammonium when glucose was the carbon source. A non-inducible mutant, nirA-1, in the regulatory gene for nitrate reductase, had high phosphoglucomutase activity when grown with nitrate or ammonium. A constitutive mutant nirAc1, in the regulatory gene for nitrate reductase had low phosphoglucomutase activity when grown with nitrate or ammonium. The mutants nir-1 and nirAc1 are recessive and semi-dominant respectively for abnormal phosphoglucomutase activity.

Aspergillus nidulans↗
PubMed1978

The acidic amino acid transport system of the baby hamster kidney cell line BHK21-C13.

The uptake of L-glutamate into BHK21-C13 cells in culture has been studied. This amino acid appears to be transported via a relatively high affinity, low capacity, Na+-dependent transport system capable of the rapid accumulation of substrate amino acids. Kinetic studies of the inhibition of L-glutamate uptake has provided information as to the substrate and the molecular configuration required for transport via the glutamate transport system. This system exhibited marked substrate specificity and was only capable of transporting L-glutamate and aspartate and certain closely related acidic amino acid analogues.

Aspartic Acid↗
PubMed1978

The transport of L-alanine by the hamster kidney cell line BHK-21-C13.

The uptake of L-alanine into BHK21-C13 cells in culture has been studied. This amino acid appears to be transported essentially via a relatively low affinity, high capacity, sodium ion dependent transport system. Inhibition studies using other amino acids or their analogues provided information about the specificity of this system. This alanine transport system was shown to exhibit a broad substrate specificity and appeared to be capable of transporting most naturally occurring neutral alpha-amino acids. Kinetic studies of the inhibition of L-alanine uptake also indicated the presence of a second neutral amino acid transport system capable of transporting this amino acid. However, it is unlikely that this second uptake system contributes greatly to L-alanine uptake. Inhibition of the uptake of L-leucine indicated that this transport system has a similar specificity to the "L"-system initially described for Ehrlich ascites carcinoma cells.

Alanine↗