Fig. 4.

X-band EPR and ENDOR spectra of GcpE (and IspH) + HMBPP/IPP. (A) ¹³C ENDOR of GcpE + [u-¹³C]-HMBPP, difference spectrum (¹³C-labeled—unlabeled). The bottom inset is the A. aeolicus IspH E126A mutant + [u-¹³C]-HMBPP (from ref. 22). (B) ³¹P ENDOR of GcpE + [u-¹³C]-HMBPP. (C) Schematic illustration of HMBPP bound to an Fe₄S₄ cluster illustrating metallacycle formation. (D) EPR of GcpE + IPP. (E) ¹³C ENDOR of GcpE + [4-¹³C]-IPP, difference spectrum (¹³C-labeled–unlabeled). (F) ³¹P ENDOR of GcpE + [4-¹³C]-IPP. EPR frequency = 9.05 GHz; ENDOR frequency = 9.66 GHz; B₀ was selected as the field where the maximum EPR signal intensity was obtained: A and B, 347.9 mT; E and F, 344.4 mT. τ-averaging (64 spectra at 8 ns step with initial τ = 200 ns) were used for collecting the Mims ENDOR spectra.