Characterization of cytochrome P450 2A4 and 2A5-catalyzed 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) metabolism.

The tobacco-specific nitrosamine, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), is a potent lung carcinogen in the A/J mouse, and is believed to be a causative agent for human lung cancer. NNK requires metabolic activation by alpha-hydroxylation to exert its carcinogenic potential. The human P450, 2A6 is a catalyst of this reaction.

N-Nitrosobenzylmethylamine hydroxylation and coumarin 7-hydroxylation: catalysis by rat esophageal microsomes and cytochrome P450 2A3 and 2A6 enzymes.

N-Nitrosobenzylmethylamine (NBzMA) is a potent and selective esophageal carcinogen in the rat and may be a causative agent for human esophageal cancer. This nitrosamine, like most, must be metabolically activated to exert its carcinogenic potential. NBzMA may be metabolized by P450-catalyzed methyl or methylene hydroxylation; the latter is believed to be the activation pathway.

Labeling of recombinant protein for NMR spectroscopy: global and specific labeling of the rat liver fructose 2,6-bisphosphatase domain.

Methods for the efficient use of the 13C-labeled nutrients, glucose and histidine, in the production of recombinant protein were developed to provide the large amount of sample required for NMR studies. The nutrient requirements were reduced by determining the minimum amount of these metabolites needed during both the growth and the induction phases of the BL21(DE3) and newly constructed BL21(DE3) histidine auxotrophic Escherichia coli cultures. These methods were developed using the separate bisphosphatase domain of rat liver 6-phosphofructo-2-kinase/ fructose-2,6-bisphosphatase, which is expressed to high levels in the pET3a/BL21 (DE3) bacterial system.