Neutron diffraction structure of (2R,3R)-L-(-)-[2-D]carnitine tetrachloroaurate, [(CH₃)₃N-CH₂-CHOH-CHD-COOH]⁺[AuCl₄]^-: Determination of the absolute stereochemistry of the crotonobetaine-to-carnitine transformation catalyzed by L-carnitine dehydratase from Escherichia coli

Single-crystal neutron diffraction has been used to determine the stereochemical course of the hydration of trans-crotonobetaine to L-(-)- carnitine by the enzyme L-carnitine dehydratase. Firstly, an X-ray analysis of the undeuterated carnitinium salt [(CH₃)₃N-CH₂-CH(OH)-CH₂- COOH]⁺[AuCl₄]^- confirmed that the absolute configuration at the C₃ position of L-(-)-carnitine (the CHOH group) is indeed R. This was accomplished using the gold atom as an anomalously-scattering source. Then stereospecifically deuterated L-(-)-[2-D] carnitine was prepared by the hydration of trans-crotonobetaine in D₂O using purified L-carnitine dehydratase from Escherichia coli. The subsequent neutron analysis of deuterated [(CH₃)₃N-CH₂-CH(OH)-CH₂-COOH]⁺ [AuCl₄]^- revealed that the CHD group at position C₂ also had the absolute R configuration, this establishing that the addition of D₂O across the C-C double bond of trans- crotonobetaine proceeds by a stereospecific syn pathway. In contrast, all other hydration-dehydration reactions examined thus far show that, when the proton added or abstracted is bonded to a carbon atom that is adjacent to a carboxylate group, the absolute stereochemistry of the resulting transformation is anti. Crystallographic details for [(CH₃)₃N-CH₂-CH(OH)- CHD-COOH]⁺[AuCl₄]^- are as follows: space group P2₁2₁2₁ (orthorhombic), a = 10.855-(2), b = 11.678(3), c = 22.776(6) Å; Z = 8; final agreement factor for the neutron analysis at 15 K: R(F(o)) = 0.097 based on 1140 reflections with 1 > 3σ(1).