Journal of Bacteriology, May 2000, p. 2544-2550, Vol. 182, No. 9
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Departments of Food Science and Technology1 and Agronomy and Range Science,2 University of California, Davis, California 95616
Received 11 June 1999/Accepted 16 February 2000
Listeria monocytogenes is a foodborne pathogen known for its tolerance to conditions of osmotic and chill stress. Accumulation of glycine betaine has been found to be important in the organism's tolerance to both of these stresses. A procedure was developed for the purification of membranes from L. monocytogenes cells in which the putative ATP-driven glycine betaine permease glycine betaine porter II (Gbu) is functional. As is the case for the L. monocytogenes sodium-driven glycine betaine uptake system (glycine betaine porter I), uptake in this vesicle system was dependent on energization by ascorbate-phenazine methosulfate. Vesicles lacking the gbu gene product had no uptake activity. Transport by this porter did not require sodium ion and could be driven only weakly by artificial gradients. Uptake rates could be manipulated under conditions not affecting secondary transport but known to affect ATPase activity. The system was shown to be both osmotically activated and cryoactivated. Under conditions of osmotic activation, the system exhibited Arrhenius-type behavior although the uptake rates were profoundly affected by the physical state of the membrane, with breaks in Arrhenius curves at approximately 10 and 18°C. In the absence of osmotic activation, the permease could be activated by decreasing temperature within the range of 15 to 4°C. Kinetic analyses of the permease at 30°C revealed Km values for glycine betaine of 1.2 and 2.9 µM with Vmax values of 2,200 and 3,700 pmol/min · mg of protein under conditions of optimal osmotic activation as mediated by KCl and sucrose, respectively.
Present address: The National Food Laboratory, Inc., Dublin, Calif.
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