Organization of Enzymes in the Polyaromatic Synthetic Pathway: Separability in Bacteria

Abstract

Ultracentrifugation in sucrose density gradients was employed to estimate the molecular weights and to determine possible physical aggregation of the five enzymes catalyzing steps two to six in the prechorismic acid portion of the polyaromatic synthetic pathway in six species of bacteria: Escherichia coli, Salmonella typhimurium, Aerobacter aerogenes, Bacillus subtilis, Pseudomonas aeruginosa, and Streptomyces coelicolor. The five enzymes were not aggregated in extracts of any of the species examined, nor are the genes encoding these enzymes clustered in those bacterial species for which genetic evidence exists. (An initial examination of the blue-green alga Anabaena variabilis indicates nonaggregation in this species also.) This situation in bacteria is in marked contrast to that found in Neurospora crassa and other fungi in which the same five enzymes are associated as an aggregate encoded (at least in the case of N. crassa) by a cluster of five genes. In addition, also in contrast to N. crassa, no evidence was obtained for more than one kind of dehydroquinase activity in any of the bacteria examined. These comparative results are discussed in relation to the origin, evolution, and functional significance of the gene-enzyme relationships existing in the early steps of aromatic biosynthesis in bacteria and fungi. other bacteria. The coliform bacteria and P. aeruginosa were also grown in minimal medium supplemented with 500 jsg of quinic acid per ml. Log-phase cells were harvested by centrifugation, washed in 0.05 M tris(hydroxymethyl)aminomethane (Tris)-hydrochlo-222

The Saccharomyces cerevisiae ARO1 gene An example of the co‐ordinate regulation of five enzymes on a single biosynthetic pathway

“…Northern blot analysis and sequence studies indicate that ARO1 is regulated by the well characterised S. cerevisiae 'general control' mechanism. This provides a very economical means of simultaneously tailoring the synthesis of five shikimate pathway enzymes to the needs of the cell.products are synthesised [4,5]. Arom genes have been cloned from a number of organisms, including S. cerevisiae [6], Aspergillus nidulans [7], Schizosaccharomyces pombe [8] and Neurospora crassa [9].We report here that in S. cerevisiae, the five shikimate pathway enzymes encoded by the ARO1 gene are co-ordinately regulated in response to amino acid limitation by the so-called 'general control' mechanism [10-13].…”

“…products are synthesised [4,5]. Arom genes have been cloned from a number of organisms, including S. cerevisiae [6], Aspergillus nidulans [7], Schizosaccharomyces pombe [8] and Neurospora crassa [9].…”

Cloning of a sequence of Aquaspirillum magnetotacticum that complements the aroD gene of Escherichia coli

“…The aroD-and the qa2-co6e6 enzymes are currently placed in separate classes of 3-dehydroquinases (Coggins and Boocock, 1986). The araD-coded enzyme of E. CO//is monofunctionai (Berlyn and Giles, 1969;Chaudhuri et ai, 1986) and catalyses the third step in the common pathway for the biosynthesis of aromatic amino acids. The qa gene cluster of N. crassa specifies the inducible enzymes involved in the utilization of quinate or shikimate as carbon source (Greever et al.…”

Isolation of a bifunctional domain from the pentafunctional arom enzyme complex of Neurospora crassa

“…The five enzyme activities which in Neurospora crassa occur on the pentafunctional arom polypeptide chain are very differently organized in some other organisms. In Escherichia coli the genes coding for the five enzyme activities are separately located (Bachmann & Low, 1980) and there is no evidence of physical association (Berlyn & Giles, 1969). In a number of yeasts (Bode & Birnbaum, 1981) and fungi (Ahmed & Giles, 1969) the five enzymes occur as a multienzyme complex, although no evidence concerning the number and kind of subunits is available.…”