The product of theade1 gene inSchizosaccharomyces pombe: A bifunctional enzyme catalysing two distinct steps in purine biosynthesis

Abstract

The assignment of the known ade genes to steps in purine biosynthesis in Schizosaccharomyces pombe has been completed with the demonstration that an ade3 mutants lacks FGAR amidotransferase, ade1A mutants lack GAR synthetase and ade1B mutants lack AIR synthetase. A comparison of enzyme activity with map position for ade1 mutants shows that (1) complementing ade1A mutants lack GAR synthetase but posses wild type amounts of AIR synthetase, (2) complementing ade1B mutants lack AIR synthetase but posses variable amounts of GAR synthetase, (3) non-complementing mutants lack both activities. In wild type strains the two activities fractionate together throughout a hundred-fold purification. Hence the ade1 gene appears to code for a bifunctional enzyme catalysing two distinct steps in purine biosynthesis. The two activities are catalysed by two different regions of the polypeptide chain which can be altered independently by mutation. Gel filtration studies on partially purified enzymes from wild type and various complementing mutant strains, indicate that the bifunctional enzyme is a multimer consisting of between four and six sub-units of 40,000 daltons each. GAR synthetase activity is associated with both the monomeric and multimeric forms but AIR synthetase is only associated with the multimer. A comparison of enzyme levels between diploids and their original complementing haploid strains suggests that complementation is due to hybrid enzyme formation.

Isolation of a multifunctional protein with aminoimidazole ribonucleotide synthetase, glycinamide ribonucleotide synthetase and glycinamide ribonucleotide transformylase activities: characterization of the aminoimidazole ribonucleotide synthetase

“…Early complementation studies using mutants of yeast deficient in purine biosynthesis in Schizosaccharomyces pombe (Fluri et al, 1976) and Saccharomyces cerevisiae (Gross & Woods, 1971) suggested that GAR synthetase and AIR synthetase activities might reside on a single bifunctional polypeptide. In fact, biochemical studies by Fluri et al in 1976 indicated that these two activities copurified =100-fold from S. pombe and suggested that both activities were confined to a polypeptide of Mr 40 000. Interestingly, GAR synthetase was active both as a monomer (Mr 40000) and in multimeric forms (Mr 160000-240000), while AIR synthetase activity could only be associated with the multimeric forms.…”

Multiple purine pathway enzyme activities are encoded at a single genetic locus in Drosophila.

“…GARS and AIRS are encoded at the same complex locus in yeasts but at a position separate from GART (26)(27)(28). The human Gart locus has been hypothesized to lie on human chromosome 14 based on complementation of the hamster Ade-E mutation by human chromosome 14 (29), whereas the loci for GARS and AIRS have been assigned to chromosome 21 (30,31), most likely in the q22 region of the chromosome (32).…”

Demonstration, by somatic cell genetics, of coordinate regulation of genes for two enzymes of purine synthesis assigned to human chromosome 21.

“…First, these have both been well-characterized biochemically (9,10). Second, coordinate regulation ofthese genes has been found for the corresponding genes in Schizosaccharomyces pombe (11). Third, we possessed a number of mutants of each complementation group.…”

“…The following 23 isozymes and 1 nutritional marker were assayed in the hybrid clones by using methods as described (19). Numbers in brackets correspond to human chromosome assignments (20): 6-phosphogluconate dehydrogenase [1]; malate dehydrogenase, soluble [2]; 1-galactosidase [3]; phosphoglucomutase 2 [4]; arylsulfatase B [5]; superoxide dismutase, mitochondrial [6]; f-glucuronidase [7]; glutathione reductase [8]; adenylate kinase [9]; proline requirement [10]; lactate dehydrogenase A [11]; lactate dehydrogenase B [12]; esterase D [13]; nucleoside phosphorylase [14]; hexosaminidase-A and pyruvate kinase [15]; phosphogluconate phosphatase [16]; galactokinase [17]; peptidase A [18]; glucose phosphate isomerase [19]; adenosine deaminase [20]; superoxide dismutase, soluble [21]; arylsulfatase-A [22]; and glucose-6-phosphate dehydrogenase [X].…”