Iron is a necessary micronutrient for organisms, it is incorporated into proteins as a biocatalyst or into electron carrier proteins vital for many biological processes. The importance of iron for bacterial survival and pathogenesis is evident by the variety of highly conserved and broadly employed iron acquisition strategies among bacterial pathogens including siderophore production, heme uptake systems, and hemophore-receptor systems. Humans and other mammals even coordinate an immune response specifically limiting iron availability to invading microorganisms through nutritional immunity. In environments where iron or heme levels are low, the ability to express high-affinity receptors specific for heme and heme degrading enzymes provide a significant advantage to the pathogen. Bacterial pathogens liberate iron from heme through heme-degrading enzymes, and the ability to utilize heme as an iron source is essential for virulence and pathogenesis. Vibrio cholerae and Escherichia coli O157:H7 are common hemolytic enteric pathogens that infect the lower intestine and have been shown to use heme as an iron source; however, the catabolic fate of heme and how iron is scavenged in an anaerobic environment remained unknown. Herein, we demonstrate that like E. coli O157:H7, V. cholerae encodes a radical S-adenosylmethionine (SAM) methyl transferase (HutW) involved in the anaerobic opening of the porphyring ring of heme. However, in contrast to the E. coli O157:H7 enzyme, there are notable differences in the genetic cluster, mechanism and products, including the ability to utilize reduced nicotinamide adenine dinucleotide phosphate (NADPH) as an electron source. The ability of HutW to use NADPH directly is unique in the radical SAM (RS) field, where most RS enzymes require either a chemical reductant or a redox partner protein to reduce the catalytic [4Fe-4S] cluster. Therefore, we subsequently pursued elucidation for this electron transfer mechanism where we showed that either the heme porphyrin ring and/or subsequent tetrapyrrole intermediates facilitate electron transfer from NADPH to the [4Fe-4S] cluster. This work not only expands the RS field, but also rationalizes the genetic cluster arrangement for enteric bacteria lacking an anaerobilin reductase.
