On the basis of biochemical studies pyruvate kinase has been identified as a key regulatory enzyme in a number of organisms, however, there is little genetic evidence to substantiate this. In Aspergillus nidulans pyruvate kinase activity is high following growth on glycolytic carbon sources and low following growth on gluconeogenic carbon sources suggesting the enzyme is heavily regulated. This seemed an excellent model system to study the role and regulation of a key enzyme of carbon metabolism and a study was therefore undertaken involving the isolation of mutants of A.nidulans lacking pyruvate kinase activity and of strains producing pyruvate kinase constitutively. Following mutagenesis and filtration enrichment a number of mutants were isolated which failed to grow on glycolytic carbon sources but grew normally on gluconeogenic carbon sources. Ten such mutants lacked pyruvate kinase activity and identify a single gene locus, pkiA (chromosome V), probably the structural gene for the enzyme. The pyruvate kinase deficient pkiA mutants, though able to grow on gluconeogenic carbon sources, are poisoned by the addition of a glycolytic carbon source such as glucose or fructose, probably due to accumulation of a toxic metabolic product. A number of fructose-insensitive revertants of the pkiA1 strain were selected in an attempt to isolate strains defective in sugar transport. Amongst these revertants, two were shown to have altered sugar uptake characteristics, demonstrating that the reversion technique described is capable of generating bona fide sugar uptake mutants in A.nidulans. One interesting observation made during this work was that the presence of agar in growth media can generate spurious results. This was observed first for a number of acetate non-utilising (acu) mutants whose growth on a number of carbon sources was impaired by agar. It was subsequently demonstrated that A.nidulans can use agar as sole carbon source by the pathway of C2 metabolism.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:468490 |
| Date | January 1978 |
| Creators | Payton, Mark Anthony |
| Publisher | University of Leicester |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://hdl.handle.net/2381/34458 |
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