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J. Bacteriol. doi:10.1128/JB.01397-06
Copyright (c) 2006, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

The lspA gene encoding the Type II Signal Peptidase of Rickettsia typhi: Transcriptional and Functional Analysis

Department of Microbiology and Immunology, University of Maryland School of Medicine, 660 West Redwood Street, Baltimore, MD 21201

^* To whom correspondence should be addressed. Email: mrahm001{at}umaryland.edu,

	Abstract

Lipoprotein processing by the type II signal peptidase (SPase II) is known to be critical for intracellular growth and virulence for many bacteria, but its role in rickettsiae is unknown. Here, we describe the analysis of lspA encoding a putative SPase II, an essential component of lipoprotein processing in gram-negative bacteria, from Rickettsia typhi. Alignment of deduced amino acid sequences shows the presence of highly conserved residues and domains that are essential for SPase II activity in lipoprotein processing. The transcription of lspA, lgt (encoding prolipoprotein transferase) and lepB (encoding type I signal peptidase) monitored by real time qRT-PCR reveals a differential expression pattern during various stages of rickettsial intracellular growth. The higher transcriptional level of all three genes at preinfection time point indicates that only live and metabolically active rickettsiae are capable of infection and inducing host cell phagocytosis. The lspA and lgt, which are involved in lipoprotein processing, show a similar level of expression. However, lepB, which is involved in non-lipoprotein secretion, shows higher level of expression, suggesting that LepB is the major signal peptidase for protein secretion and supports our in silico prediction that out of 89 secretory proteins, only 14 are lipoproteins. Overexpression of R. typhi lspA in Escherichia coli confers increased globomycin resistance, indicating its function as SPase II. In genetic complementation, the recombinant lspA from R. typhi significantly restores the growth of temperature sensitive E. coli Y815 at non-permissive temperature, supporting its biological activity as SPase II in prolipoprotein processing.

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