The characterisation of transketolase from Leishmania mexicana

Veitch, Nicola Jean (2002) The characterisation of transketolase from Leishmania mexicana. PhD thesis, University of Glasgow.

Abstract

Transketolase is a key enzyme of the non-oxidative branch of the pentose phosphate pathway. The identification and characterisation of transketolase in Leishmania mexicana will aid in the understanding of the specific interactions of this enzyme and its products within this biochemical pathway. The pentose phosphate pathway may be a potential target for chemotherapy against Leishmania as it provides several key intermediates necessary for parasite survival within the host cell. In this study, the transketolase gene from L. mexicana was identified and cloned using a variety of molecular methodologies. The transketolase was shown to be similar to transketolases from other organisms, and contained two conserved motifs, a thiamine-pyrophosphate binding motif and a transketolase motif Analysis of the transketolase sequence revealed a carboxy-terminal peroxisome-targeting signal (PTS), suggestive of a glycosomal subcellular localisation. Subcellular fractionation of the promastigote Leishmania revealed that the protein was predominantly cytosolic, however a glycosomal localisation cannot be ruled out in other life cycle stages. Putative L. major and T. brucei transketolase protein sequences also contain a PTS at the C-terminal of the protein. The cloned transketolase gene was expressed as a recombinant protein using a bacterial system with the enzyme containing an N-terminal histidiae tag, allowing for efficient purification. Overexpression of the soluble transketolase protein occurred only at a low temperature (15

• Item Type: Thesis (PhD)
• Qualification Level: Doctoral
• Keywords: Biochemistry.
• Colleges/Schools: College of Medical Veterinary and Life Sciences
• Supervisor's Name: Barrett, Mike
• Date of Award: 2002
• Depositing User: Enlighten Team
• Unique ID: glathesis:2002-72313
• Copyright: Copyright of this thesis is held by the author.
• Date Deposited: 24 May 2019 15:12
• Last Modified: 08 Sep 2022 10:33
• Thesis DOI: 10.5525/gla.thesis.72313
• URI: https://theses.gla.ac.uk/id/eprint/72313