Robson, Paul R. H. (1991) Towards the mechanism of carotenogenesis in Myxococcus xanthus. PhD thesis, University of Warwick.

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Abstract

Myxococcus xanthus Is a Gram negative, heterotrophic, soil dwelling, bacterium. It exhibits a number of interesting characteristics. Including the synthesis of carotenoids. Carotenoids protect the cells from lethal photolysis by protoporphyrin IX In the presence of light (Burchard and Dworking, 1966, J.Bac., 97, 1165-1168). The region of DNA which encodes the proteins responsible for the control of carotenogeneals by light has been identified (Martinez-Laborda and Murillo, 1989, Genetics, 122, 481-490, Hodgson, manuscript in preparation). This region, termed oafQRS, is under the control of a light inducible promoter, termed pQRS. The alma of study were to determine the mechanism by which light regulates pQRS, and to establish its possible application to biotechnology by reconstituting the light inducible mechanism in a heterologous host, Escherichia coll. To facilitate these studies the pQRS promoter was linked to a promoter-less lacZ gene, allowing a simple assay for promoter activity.

The pQRS region exhibits two expression patterns In E. coll, which depend on the size of the pQRS region studied. Only one of these is believed to be the expression of pQRS, showing a low level of expression which may be increased in the presence of the carQRS region. The second expression pattern is believed due to a promoter-like region upstream of pQRS, term pX. A E. coli strain permeable to protoporphyrin IX has been isolated. This strain exhibits photolysis, in the presence of protoporphyrin IX, which may be quenched by carotenoids, and thus is an ideal system for further studies of the Induction of pQRS in E. coll.

The pQRs promoter has been extensively studied in its native host.
Genetic studies in which a mutant of M. xanthus deleted for carQRS was
reconstituted with fragments of the region showed an open reading frame previously without function was required for full expression from pQRS .

Light Induction of batch cultures of M. xanthus was used to study in part the physiology of the inductive process. The level to which pQRS activity is induced by light decreases throughout the growth cycle, and is subject to feedback control by endogenous carotenoids. The pQRS promoter may be induced by defined laser light of wavelength 410nm, the maxima of absorption by protoporphyrin IX. Extensive continuous culture experiments showed that the pQRS promoter may be induced in the dark by increasing the oxygenation of the culture. Typically two-fold induction was seen. In addition it was shown that the level of oxygenation determined the extent to which light stimulates expression from pQRS.

The reactive species involved in the stimulation of pQRS activity by light and oxygen was investigated. The promoter could not be induced by exogenous hydrogen peroxide, or by the presence of methyl viologen, a superoxide radical generator. However, it was shown that the photoactive dye, Toluldlne Blue O, could induce expression from pQRS under red light stimulation which does not stimulate the native photoreceptor. This Induction was shown to be due to singlet oxygen by a singlet oxygen quencher. Additionally it was shown that Induction of pQRS by the native photoreceptor could be quenched in the same way. Thus It has been shown that light Induces carotenogenesis in M. xanthus through singlet oxygen.

Item Type:	Thesis or Dissertation (PhD)
Subjects:	Q Science > QK Botany Q Science > QP Physiology Q Science > QR Microbiology
Library of Congress Subject Headings (LCSH):	Myxococcus xanthus, Carotenoids
Official Date:	November 1991
Dates:	Date Event November 1991 UNSPECIFIED
Institution:	University of Warwick
Theses Department:	Department of Biological Sciences
Thesis Type:	PhD
Publication Status:	Unpublished
Supervisor(s)/Advisor:	Hodgson, D. A. (David A.)
Format of File:	pdf
Extent:	xxiv, 221 leaves : illustrations
Language:	eng

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