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J. Bacteriol. doi:10.1128/JB.00057-08
Copyright (c) 2008, American Society for Microbiology and/or the Listed Authors/Institutions. All Rights Reserved.

Purification and Properties of the Plasmid Maintenance Proteins from the B. burgdorferi Linear Plasmid lp17

Department of Biochemistry & Molecular Biology and Department of Microbiology & Infectious Diseases, University of Calgary, 3330 Hospital Drive N.W., Calgary, AB T2N 4N1, CANADA

^* To whom correspondence should be addressed. Email: chaconas{at}ucalgary.ca.

	Abstract

The Lyme disease spirochete Borrelia burgdorferi carries more plasmids than any other bacterium, many of which are linear with covalently closed hairpin ends. These plasmids have also been referred to as mini-chromosomes and essential genetic elements and are integral components of its segmented genome. We have investigated two plasmid maintenance proteins, BBD14 (the replication initiator) and BBD21 (a presumptive ParA orthologue), encoded by the linear plasmid lp17; these proteins are representatives of paralogous families 62 and 32, respectively. We have purified recombinant 6-his-BBD21 and shown it to possess an ATPase activity. 6-his-BBD14 initially could not be overexpressed in E. coli by itself. It was only effectively overproduced in recombinant form through co-expression with other B. burgdorferi proteins and codon optimization. Although the mechanism for increased production through co-expression is not clear, this method holds promise for expression and purification of other B. burgdorferi proteins, a number of which have remained recalcitrant to purification from E. coli. Finally, we present evidence for physical interaction of BBD14 and BBD21, a feature suggesting that BBD21 and the paralogous family 32 proteins are more likely involved in DNA replication rather than functioning as simple ParA orthologues as previously surmised based upon sequence homology. Such a role would not preclude a function in plasmid partitioning through interaction with the replication initiator.