Enantiomeric Free Radicals and Enzymatic Control of Stereochemistry in a Radical Mechanism:  The Case of Lysine 2,3-Aminomutases

Abstract

The product of yjeK in Escherichia coli is a homologue of lysine 2,3-aminomutase (LAM) from Clostridium subterminale SB4, and both enzymes catalyze the isomerization of (S)- but not (R)-alpha-lysine by radical mechanisms. The turnover number for LAM from E. coli is 5.0 min(-1), 0.1% of the value for clostridial LAM. The reaction of E. coli LAM with (S)-alpha-[3,3,4,4,5,5,6,6-(2)H8]lysine proceeds with a kinetic isotope effect (kH/kD) of 1.4, suggesting that hydrogen transfer is not rate-limiting. The product of the E. coli enzyme is (R)-beta-lysine, the enantiomer of the clostridial product. Beta-lysine-related radicals are observed in the reactions of both enzymes by electron paramagnetic resonance (EPR). The radical in the reaction of clostridial LAM has the (S)-configuration, whereas that in the reaction of E. coli LAM has the (R)-configuration. Moreover, the conformations of the beta-lysine-related radicals at the active sites of E. coli and clostridial LAM are different. The nuclear hyperfine splitting between the C3 hydrogen and the unpaired electron at C2 shows the dihedral angle to be 6 degrees, unlike the value of 77 degrees reported for the analogous radical bound to the clostridial enzyme. Reaction of (S)-4-thialysine produces a substrate-related radical in the steady state of E. coli LAM, as in the action of the clostridial enzyme. While (S)-beta-lysine is not a substrate for E. coli LAM, it undergoes hydrogen abstraction to form an (S)-beta-lysine-related radical with the same stereochemistry of hydrogen transfer from C2 of (S)-beta-lysine to the 5'-deoxyadenosyl radical as in the action of the clostridial enzyme. The resulting beta-lysyl radical has a conformation different from that at the active site of clostridial LAM. All evidence indicates that the opposite stereochemistry displayed by E. coli LAM is determined by the conformation of the lysine side chain in the active site. Stereochemical models for the actions of LAM from C. subterminale and E. coli are presented.

Loss of Elongation Factor P Disrupts Bacterial Outer Membrane Integrity

“…Two other laboratories have independently corroborated aspects of these findings (6,60). YjeK belongs to the family of 2,3-lysine aminomutases, ironsulfur cluster-containing enzymes that catalyze the interconversion of L-lysine and 3,6-diaminohexanoic acid (also known as ␤-lysine) (8). While PoxA bears close homology to the catalytic domain of the class II lysyl-tRNA synthetase (LysRS) family of enzymes that catalyze the addition of lysine to its cognate tRNA Lys , a number of studies have failed to show that PoxA can aminoacylate a tRNA (1,36,37).…”

PoxA, YjeK, and Elongation Factor P Coordinately Modulate Virulence and Drug Resistance in Salmonella enterica

“…These iron-sulfur cluster-containing enzymes catalyze the transfer of the alpha-carbon amino group of lysine to the β-carbon to generate the product β-lysine. Lysine aminomutases display stereospecificity and YjeK belongs to the group that catalyze the conversion of L -lysine to (R)-β-lysine (Behshad et al, 2006). No biological function for this enzyme has been ascribed thus far in either E. coli or Salmonella , although in other bacterial species β-lysine can act as a compatible solute to protect the cell from osmotic stress or as an intermediate for the generation of secondary metabolites (Muller et al, 2005).…”

Elongation factor P: Function and effects on bacterial fitness

“…YjeK is a homolog of lysine 2, 3 aminomutase (LAM), but it lacks the multimerization domain found in that protein family. While LAM proteins usually promote the conversion of S‐α‐Lys to S‐β‐Lys, E. coli YjeK converts S‐α‐Lys to R‐β‐Lys, which is the first step on the EF‐P modification pathway. YjeA is a paralog of the lysyl‐tRNA synthetases (Lys‐RS2), which charge tRNA Lys with Lys.…”