Cloning of the Pneumocystis jirovecii trifunctional FAS gene and complementation of its DHPS activity in Escherichia coli
- Author(s)
- Iliades, P; Walker, DJ; Castelli, L; Satchell, J; Meshnick, SR; Macreadie, IG;
- Details
- Publication Year 2004-12,Volume 41,Issue #12,Page 1053-1062
- Journal Title
- FUNGAL GENETICS AND BIOLOGY
- Publication Type
- Journal Article
- Abstract
- Pneurnocystis pneumonia or PCP is caused by Pneumocystisjirovecii, an obligate parasite of the human lung. In this study P. jirovecii genomic sequence encoding FAS, a trifunctional protein including dihydroneopterin aldolase (DHNA), hydroxymethyldihydropterin pyrophosphokinase (PPPK) and dihydropteroate synthase (DHPS) were identified by PCR amplification from fixed broncheolar lavage samples from patients having Pneumocystis pneumonia. The P. jirovecii trifunctional DHNAPPPK-DHPS genes (PjFAS) showed a high degree of conservation with the rat Pneurnocystis carinii and P. earimif sp. macaca sequences. To test the functionality of the PjFAS sequences introns were removed followed by cloning and expression of PjFAS sequences in a DHPS-disrupted Escherichia coli strain. Complementation depended on the presence of N-terminal FAS sequences in addition to a glutathione S- transferase tag to the N-terminus of PjFAS. Functional complementation allowed evaluation of DHPS mutations implicated with sulfa drug resistance. (C) 2004 Elsevier Inc. All rights reserved.
- Publisher
- ACADEMIC PRESS INC ELSEVIER SCIENCE
- Keywords
- DIHYDROPTEROATE SYNTHASE GENE; HUMAN-IMMUNODEFICIENCY-VIRUS; SULFONE PROPHYLAXIS FAILURES; SACCHAROMYCES-CEREVISIAE; CARINII-PNEUMONIA; HYDROXYMETHYLDIHYDROPTERIN PYROPHOSPHOKINASE; DIHYDRONEOPTERIN ALDOLASE; CRYSTAL-STRUCTURE; AIDS PATIENTS; MUTATIONS
- Publisher's Version
- https://doi.org/10.1016/j.fgb.2004.08.006
- Terms of Use/Rights Notice
- Refer to copyright notice on published article.
Creation Date: 2004-12-01 12:00:00