Analysis of cloned genes for GABA catabolism.
thesisposted on 2015-11-19, 09:07 authored by Sudjadi.
A putative hpa+ clone has been isolated allowing an Hpa-Hpc+ mutant CD03 to growth on 4-HPA. Detection of a 4-HPA monooxygenase from extracts was unsuccessful. When it became apparent that gab genes had been cloned the emphasis switched to detailed analysis of those genes. The gab genes were isolated using suppression of a mutant CT101 defective in the sad gene. It seems that the cloned gabDT genes encoding NADP-dependent SSA dehydrogenase and GABA transaminase are responsible for these, not the gabD gene only. Possible explanation of these phenomena are described. E.coli C and K-12 cannot utilise GABA as the sole carbon source. However, E.coli K-12 can mutate to grow on GABA at 30°C but did not at 37°C without further mutation. Both types of mutants were found to have high activities of GABA utilizing enzymes. Studying clones isolated from the wild-type and mutated clones revealed there was a negative regulatory gene, gabR. E.coli K-12 can utilise GABA as the sole nitrogen source at 30°C but not at 37°C. The 1.7 kbp EcoRI-SalI fragment from the wild-type and mutated clones allowed cells to utilise GABA as the sole nitrogen source at 37°C. Thus, the fragment might encode a positive regulatory gene called "gabC". The gene order was determined to be gab"C"DTPR and there were located within the 10.1 kbp EcoRI region. Transcription of gabDTP is from the gabD to gabP direction, whereas gabR appears to be transcribed in the opposite direction. A representation of the organisation and expression of gab genes in E.coli is proposed.
Date of award1991-01-01
Awarding institutionUniversity of Leicester