Abstract
S-adenosyl-l-methionine (SAM) is a crucial enzymatic cofactor that is conserved across all domains of life. Despite the pivotal role of this cofactor, its chirality at the sulfonium sulfur and the differing biological activities of its epimers, (S,S)-SAM and (R,S)-SAM, are often overlooked. Although enzymes predominantly utilize the (S,S)-SAM epimer, due to spontaneous epimerization at the sulfonium sulfur of SAM, the (R,S)-SAM epimer is present in all cells as well as in commercial SAM-containing products. Recently, an enzyme containing the DUF62 domain, identified as Salinispora tropica (StDUF62), has been shown to selectively hydrolyze (R,S)-SAM. It has been hypothesized that this function prevents the problematic accumulation of this epimer. Fluorinases, the only enzymes known to naturally incorporate fluorine into organic compounds, are homologous to enzymes of the DUF62 family. The discovery of unexpected diastereoselectivity of StDUF62 however raised an important question regarding the diastereoselectivity of the evolutionarily related bacterial fluorinase FlA1, an enzyme of significant importance. Given the relationship between these enzymes and their similar catalytic functions, it would be reasonable to hypothesize that FlA1 might also demonstrate activity toward the (R,S)-SAM diastereomer. Despite this homology, we report here the opposite diastereoselectivity of StDUF62 and Streptomyces sp. MA37 fluorinase (FlA1). The unusual lysine-free amino acid composition of StDUF62 suggests an evolutionary origin in haloadaptation; however, its SAM-hydrolyzing activity is greatly diminished at physiological concentrations of KCl or NaCl. We show that this inhibition is not caused solely by the competition with the chloride anion, as Na(2)SO(4) at equivalent ionic strength is also greatly diminishing StDUF62 activity, contrary to the fluorinating activity of FlA1. Both adenosine and increased ionic strength promoted StDUF62 trimer formation, whereas increased ionic strength alone led to inhibition. Considering the contrast between the wasteful hydrolysis of (R,S)-SAM and the energetically efficient mechanisms of eukaryotic (R,S)-SAM recycling, we suggest that (R,S)-SAM hydrolysis might not be the physiological function of StDUF62.