Temperature-Induced Changes in the Lipopolysaccharide ofYersinia pestisAffect Plasminogen Activation by the Pla Surface Protease

Abstract

The Pla surface protease ofYersinia pestisactivates human plasminogen and is a central virulence factor in bubonic and pneumonic plague. Pla is a transmembrane β-barrel protein and member of the omptin family of outer membrane proteases which require bound lipopolysaccharide (LPS) to be proteolytically active. Plasminogen activation and autoprocessing of Pla were dramatically higher inY. pestiscells grown at 37°C than in cells grown at 20°C; the difference in enzymatic activity by far exceeded the increase in the cellular content of the Pla protein.Y. pestismodifies its LPS structure in response to growth temperature. We purified His₆-Pla under denaturing conditions and compared various LPS types for their capacity to enhance plasmin formation by His₆-Pla solubilized in detergent. Reactivation of His₆-Pla was higher withY. pestisLPSs isolated from bacteria grown at 37°C than with LPSs from cells grown at 25°C. Lack of O antigens and the presence of the outer core region as well as a lowered level of acylation in LPS were found to enhance the Pla-LPS interaction. Genetic substitution of arginine 138, which is part of a three-dimensional protein motif for binding to lipid A phosphates, decreased both the enzymatic activity of His₆-Pla and the amount of Pla inY. pestiscells, suggesting the importance of the Pla-lipid A phosphate interaction. The temperature-induced changes in LPS are known to helpY. pestisto avoid innate immune responses, and our results strongly suggest that they also potentiate Pla-mediated proteolysis.