Crystallization of Mycobacterium tuberculosis type I signal peptidase

Date created: 
2015-03-25
Identifier: 
etd8922
Keywords: 
Mycobacterium
Tuberculosis
Protein translocation
Signal peptidase
Ser/Lys protease
Protein crystallization
Protein crystallography analysis
Abstract: 

Bacterial signal peptidase I (SPase I) plays a key role in the translocation of proteins across the cytoplasmic membrane. SPase I cleaves the signal peptide from the pre-protein. The unique Ser/Lys dyad mechanism, utilized by SPase I, is conserved in this family of enzymes in both Gram-positive and Gram-negative species. Mycobacterium tuberculosis (Mtb) SPase I is an attractive target for the design of antimicrobial compounds since it is essential for the bacterium’s viability and is involved in the translocation of important pathogenic proteins to their final destinations. Based on sequence alignments of bacterial SPase I, it is hypothesized that Lys174 and Ser96 are catalytically active residues and act as a general base and nucleophile in the Ser/Lys dyad, respectively. We have investigated the overexpression, purification and crystallization of three full length constructs including WT Mtb SPase I, Mtb SPase I K174A and Mtb SPase I S96A. After obtaining initial protein crystals of Mtb SPase I, different optimizing methods were applied to improve the crystal quality. The optimized crystal diffracted to 3.5 Å resolution. We present here analysis of the Mtb SPase I K174A crystals and initial data analysis.

Document type: 
Thesis
Rights: 
Copyright remains with the author. The author granted permission for the file to be printed, but not for the text to be copied and pasted.
File(s): 
Supervisor(s): 
Dr. Mark Paetzel
Department: 
Science:
Thesis type: 
(Thesis) M.Sc.
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