Lysine 2,3-aminomutase (LAM) utilizes a [4Fe-4S] cluster, S-adenosyl-L-methionine (SAM), and pyridoxal 5'-phosphate (PLP) to isomerize L-alpha-lysine to L-beta-lysine. LAM is a member of the radical-SAM enzyme superfamily in which a [4Fe-4S](+) cluster reductively cleaves SAM to produce the 5'-deoxyadenosyl radical, which abstracts an H-atom from substrate to form 5'-deoxyadenosine (5'-Ado) and the alpha-Lys(center dot) radical (state 3 (Lys(center dot))). This radical isomerizes to the beta-Lys(center dot) radical (state 4(Lys(center dot))), which then abstracts an H-atom from 5'-Ado to form beta-lysine and the 5'-deoxyadenosyl radical; the latter then regenerates SAM. We use C-13, H-1,H-2, P-31, and N-14 ENDOR to characterize the active site of LAM in intermediate states that contain the isomeric substrate radicals or analogues. With L-alpha-lysine as substrate, we monitor the state with beta-Lys(center dot). In parallel, we use two substrate analogues that generate stable analogues of the alpha-Lys(center dot) radical: trans-4,5-dehydro-L-lysine (DHLys) and 4-thia-L-lysine (SLys). This first glimpse of the motions of active-site components during catalytic turnover suggests a possible major movement of PLP during catalysis. However, the principal focus of this work is on the relative positions of the carbons involved in H-atom transfer. We conclude that the active site facilitates hydrogen atom transfer by enforcing van der Waals contact between radicals and their reacting partners. This constraint enables the enzyme to minimize and even eliminate side reactions of highly reactive species such as the 5'-deoxyadensosyl radical.