Biomedical subjects

W A Frazier

Publications and source records attributed to W A Frazier.

93 records · Page 6Linked to original sources

PubMed1970

Formation of assimilatory nitrate reductase by in vitro inter-cistronic complementation in Neurospora crassa.

In vitro complementation of the soluble assimilatory nicotinamide adenine dinucleotide phosphate, reduced form (NADPH)-nitrate reductase was attained by mixing cell-free preparations of certain Neurospora nitrate reductase mutants: induced nit-1 (uniquely possessing inducible NADPH-cytochrome c reductase) with (a) uninduced or induced nit-2 or nit-3, or (b) uninduced wild type. The complementing activity of induced nit-1 is soluble while that of nit-2, nit-3, and wild type is particulate but not of mitochondrial origin. All fractions are inactivated by heat or trypsin. The NADPH-nitrate reductase enzymes formed in the above three complementing mixtures are similar to the wild-type enzyme in sucrose density gradient profiles, molecular weight, substrate affinity, sensitivity to inhibitors and temperature, but show different ratios of associated enzyme activities. The data suggest that nitrate reductase consists of at least two protein subunits: a nitrate-inductible subunit as reflected by inductible NADPH-cytochrome c reductase, and a constitutive protein which is activated (as indicated by the appearance of flavine adenine dinucleotide, reduced form (FADH(2))- and reduced methyl viologen-nitrate reductase activities) when it combines with the inductible subunit.

Cell-Free System↗

PubMed1970

In vitro assembly of Neurospora assimilatory nitrate reductase from protein subunits of a Neurospora mutant and the xanthine oxidizing or aldehyde oxidase systems of higher animals.

In vitro assembly or complementation of a hybrid assimilatory nitrate reductase was attained by mixing a preparation of nitrate-induced N. crassa mutant nit-1 specifically with acid-treated (pH 2.5) bovine milk or intestinal xanthine oxidase, rabbit liver aldehyde oxidase, or chicken liver xanthine dehydrogenase. The complementation reaction specifically required induced nit-1, the only nitrate reductase mutant of Neurospora that lacked xanthine dehydrogenase and was unable to use hypoxathine or nitrate as a sole nitrogen source. The complementing activities of the above acid-treated enzymes correspond to their xanthine or aldehyde oxidizing activity profiles on sucrose density gradients. The resulting soluble, reduced nicotinamide adenine dinucleotide phosphate (NADPH)-nitrate reductases are the same as the Neurospora wild type enzyme in sucrose density gradient profile, molecular weight, substrate affinities, and sensitivity to inhibitors and temperature. By analogy to a similar in vitro complementation of nitrate reductase in mixtures of induced nit-1 and individual nonalleic Neurospora mutants, or uninduced wild type, the complemented nitrate apparently consists of an inducible protein subunit (possessing inducible NADPH-cytochrome c reductase) furnished by nit-1 and a subunit from the acid-treated xanthine or aldehyde oxidizing system which can substitute for the constitutive component furnished by the other mutants or uninduced wild type. The data suggest that Neurospora nitrate reductase and the xanthine oxidizing system and aldehyde oxidase of animals, all of which are molybdenum-containing enzymes catalyzing the reduction of nitrate to nitrite, share a highly similar protein subunit.

Aldehydes↗

PubMedDate not supplied

Thrombospondin binds falciparum malaria parasitized erythrocytes and may mediate cytoadherence.

Plasmodium falciparum infected erythrocytes containing mature trophozoites and schizonts sequester along venular endothelium and are not in the peripheral circulation of patients with malaria. Knobs appear on infected erythrocytes and are the points of attachment to endothelium. Sequestration may protect the parasite from splenic destruction and may play a role in the pathogenesis of cerebral malaria. Correlates of sequestration have been developed in vitro using cultured human endothelium and an amelanotic melanoma cell line. Knobless strains (K-) of P. falciparum fail to sequester in vivo and to bind to cells in vitro. We now present evidence that the receptor for cytoadherence is the glycoprotein, thrombospondin. Aotus monkey or human erythrocytes containing knobby (K+) but not Aotus erythrocytes containing knobless strains of P. falciparum bind to immobilized thrombospondin. Neither binds to the adhesive proteins laminin, fibronectin, factor VIII/von Willebrand factor or vitronectin. Both soluble thrombospondin and anti-thrombospondin antibodies inhibit binding of parasitized Aotus erythrocytes to immobilize thrombospondin and to melanoma cells which secrete thrombospondin.

Animals↗