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dc.contributor.advisorSmit, M. S.
dc.contributor.advisorAlbertyn, J.
dc.contributor.authorMatatiele, Puleng Rose
dc.date.accessioned2015-11-12T12:40:54Z
dc.date.available2015-11-12T12:40:54Z
dc.date.copyright2005-05
dc.date.issued2005-05
dc.date.submitted2005-05
dc.identifier.urihttp://hdl.handle.net/11660/1621
dc.description.abstractThe cytochrome P-450 monooxygenase and b-oxidation systems of alkane-utilizing yeasts have been studied extensively, whereas very little is known about the fatty (long chain) alcohol and fatty aldehyde oxidizing enzymes. With the recent completion of sequencing of the genome of Yarrowia lipolytica, an alkane-degrading yeast, several putative aldehyde dehydrogenases (ALDHs) have been identified. Four of these were identified as fatty ALDHs (FALDHs). Northern blot analysis and RT-PCR showed that one of the FALDH genes, labelled FALDH4, is induced during growth of Y. lipolytica on alkanes, whereas another aldehyde dehydrogenase gene, labelled ALDH1, was constitutively expressed. Functional analysis of the four FALDH isogenes was initiated by single gene deletion of the four fatty aldehyde dehydrogenase isogenes in all possible combinations. The Cre-loxP recyclable tools system was used for gene disruption. Growth properties of the triple and quadruple deletion strains on alkanes were investigated. A slightly arrested growth in hexadecane was observed in two strains, the triple deletion mutant with intact FALDH2 isogene and the quadruple deletion mutant with all four FALDH isogenes deleted. Very strong hydrophobicity during growth of these mutants in hexadecane was also observed. At this stage one can only say that disruption of FALDH isogenes had a slight negative effect on growth of this yeast on alkanes.; However, it is not yet clear which individual isogenes are the most important for alkane metabolism in this organism. Although fatty aldehyde dehydrogenase (FALDH) activity has been detected in fungi no FALDH genes have yet been cloned, sequenced and expressed. Through BLAST searches using the human FALDH sequence as query we have identified 28 FALDH/FALDH-like gene sequences of which nine are from molds and 19 from yeast species. A comparative study of these sequences showed that fungal FALDH sequences may fall into several different subclasses of the ALDH3 family. Unique features of these proteins included presence of several transmembrane domains and in particular relatively long C- and N-termini. Searches of the sequenced Y. lipolytica genome for fatty alcohol oxidase (FAOD) and fatty alcohol dehydrogenase (FADH) encoding genes, which could be involved in the oxidation of fatty alcohols to aldehydes, yielded only one putative FADH encoding gene. However, FADH activity during growth on n-alkanes was very low and Northern-blot analyses showed that this gene was only eakly expressed during growth on hydrocarbon and non-hydrocarbon substrates.en_ZA
dc.description.sponsorshipNational Research Foundation (NRF)en-ZA
dc.description.sponsorshipDepartment of Labour (DoL)en_ZA
dc.language.isoenen_ZA
dc.publisherUniversity of the Free Stateen_ZA
dc.subjectThesis (Ph.D. (Microbial, Biochemical and Food Biotechnology))--University of the Free State, 2005en_ZA
dc.subjectYeast fungien_ZA
dc.subjectAlcohol dehydrogenaseen_ZA
dc.subjectEnzymesen_ZA
dc.subjectFatty alcohol dehydrogenaseen_ZA
dc.subjectFatty aldehyde dehydrogenaseen_ZA
dc.subjectRT-PCRen_ZA
dc.subjectNorthern bloten_ZA
dc.subjectSouthern bloten_ZA
dc.subjectGene disruptionsen_ZA
dc.subjectCre-loxPen_ZA
dc.subjectFungal FALDHsen_ZA
dc.subjectAlkanesen_ZA
dc.subjectYarrowia lipolyticaen_ZA
dc.titleFatty alchohol and fatty aldehyde dehydrogenases of Yarrowia lipolyticaen_ZA
dc.typeThesisen_ZA
dc.rights.holderUniversity of the Free Stateen_ZA


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