Radical S-adenosyl-L-methionine (SAM) enzymes comprise a vast superfamily catalyzing diverse reactions essential to all life through homolytic SAM cleavage to liberate the highly reactive 5 '-deoxyadenosyl radical (5 '-dAdo-). Our recent observation of a catalytically competent organometallic intermediate Omega that forms during reaction of the radical SAM (RS) enzyme pyruvate formate-lyase activating-enzyme (PFL-AE) was therefore quite surprising, and led to the question of its broad relevance in the superfamily. We now show that Omega in PFL-AE forms as an intermediate under a variety of mixing order conditions, suggesting it is central to catalysis in this enzyme. We further demonstrate that Omega forms in a suite of RS enzymes chosen to span the totality of superfamily reaction types, implicating Omega as essential in catalysis across the RS superfamily. Finally, EPR and electron nuclear double resonance spectroscopy establish that Omega involves an Fe-05 ' bond between 5 '-dAdo" and the [4Fe-4S] cluster. An analogous organometallic bond is found in the well-known adenosylcobalamin (coenzyme B-12) cofactor used to initiate radical reactions via a 5 ' dAdo" intermediate. Liberation of a reactive 5 '-dAdo" intermediate via homolytic metal-carbon bond cleavage thus appears to be similar for Omega and coenzyme B-12. However, coenzyme B-12 is involved in enzymes catalyzing only a small number (similar to 12) of distinct reactions, whereas the RS superfamily has more than 100 000 distinct sequences and over 80 reaction types characterized to date. The appearance of Omega across the RS superfamily therefore dramatically enlarges the sphere of bio-organometallic chemistry in Nature.