Endogenous superoxide is a key effector of the oxygen sensitivity of a model obligate anaerobe

Abstract

It has been unclear whether superoxide and/or hydrogen peroxide play important roles in the phenomenon of obligate anaerobiosis. This question was explored usingBacteroides thetaiotaomicron, a major fermentative bacterium in the human gastrointestinal tract. Aeration inactivated two enzyme families—[4Fe-4S] dehydratases and nonredox mononuclear iron enzymes—whose homologs, in contrast, remain active in aerobicEscherichia coli. Inactivation-rate measurements of one such enzyme,B. thetaiotaomicronfumarase, showed that it is no more intrinsically sensitive to oxidants than is anE. colifumarase. Indeed, when theE. colienzymes were expressed inB. thetaiotaomicron, they no longer could tolerate aeration; conversely, theB. thetaiotaomicronenzymes maintained full activity when expressed in aerobicE. coli. Thus, the aerobic inactivation of theB. thetaiotaomicronenzymes is a feature of their intracellular environment rather than of the enzymes themselves.B. thetaiotaomicronpossesses superoxide dismutase and peroxidases, and it can repair damaged enzymes. However, measurements confirmed that the rate of reactive oxygen species production inside aeratedB. thetaiotaomicronis far higher than inE. coli. Analysis of the damaged enzymes recovered from aeratedB. thetaiotaomicronsuggested that they had been inactivated by superoxide rather than by hydrogen peroxide. Accordingly, overproduction of superoxide dismutase substantially protected the enzymes from aeration. We conclude that when this anaerobe encounters oxygen, its internal superoxide levels rise high enough to inactivate key catabolic and biosynthetic enzymes. Superoxide thus comprises a major element of the oxygen sensitivity of this anaerobe. The extent to which molecular oxygen exerts additional direct effects remains to be determined.