Growth inhibition of Bacteroides fragilis by hemopexin: proteolytic degradation of hemopexin to overcome heme limitation

Abstract

The stimulatory effect of heme on growth of Bacteroides fragilis, an anaerobic human pathogen, was strongly inhibited by hemopexin, an avid (K d 6 1 pM) heme-binding plasma protein. Both rabbit and human hemopexins were bacteriostatic for a limited period of time, suggesting an adaptation by B. fragilis to heme-limited growth, and that hemopexin-bound heme can eventually be utilized by the bacteria. The inhibitory effect of hemopexin was lost when heme in the medium was replaced by protoporphyrin IX, which is bound less strongly by hemopexin (K d V1 WM). Protease activity was detected in the culture supernatant of B. fragilis grown in the presence of heme plus hemopexin but not in the presence of free heme, protoporphyrin IX or protoporphyrin IX plus hemopexin, suggesting that the enzyme(s) is induced by heme macrocycle limitation due to the scavenging effect of hemopexin. This protease activity was able to degrade rabbit hemopexin and human hemopexin, as well as human transferrin and ovalbumin, and may be a due to a serine protease since it was inhibited by phenylmethylsulfonyl fluoride (PMSF) but not by EDTA, leupeptin, pepstatin A or aprotinin. Thus, B. fragilis may overcome hemopexin-mediated heme limitation by secreting inducible protease(s), shown here to make protein-bound heme available to the microorganism. ß

The Medically Important Bacteroides spp. in Health and Disease

“…Apparently B. fragilis does not take up iron through the production of siderophores , but it can utilize iron from iron-bound transferrin, heme, hemoglobin and haptoglobin-hemoglobin complex Otto et al, 1994). Furthermore, B. fragilis is able to overcome hemopexin-mediated heme limitation by degradation of hemopexin, a heme-binding plasma protein, to make the protein-bound heme available for uptake (Rocha et al, 2001). This ability may enable B. fragilis to overcome the inhibitory effect of serum, which is commonly seen with many of the other Bacteroides species and can be overcome by the addition of hemin or protoporphyrin IX plus iron (Verweijvan Vught et al, 1986b).…”