Single-crystal neutron di
ffraction has been used to determine thestereochemical course o
f the hydrationo
f trans-crotonobetaine to
L-(-)-carnitine bythe enzyme
L-carnitine dehydratase. Firstly, an X-rayanalysis o
f theundeuterated carnitinium salt[(CH
3)
3N-CH
2-CH(OH)-CH
2-COOH]
+[AuCl
4]
-con
firmed that the absolute con
figurationat the C
3 position o
f L-(-)-carnitine (theCHOH group) is indeed
R. This was accomplished usingthe gold atom asan anomalously-scattering source. Then stereospeci
ficallydeuterated
L-(-)-[2-D] carnitine was prepared bythehydration o
f trans-crotonobetaine in D
2O usingpuri
fied
L-carnitine dehydratase
from
Escherichiacoli. The subsequentneutron analysis o
f deuterated[(CH
3)
3N-CH
2-CH(OH)-CH
2-COOH]
+[AuCl
4]
- revealed that the CHD group atpositionC
2 also had the absolute
R con
figuration, thusestablishing that
the addition ofD2O across the C=Cdouble bond oftrans-crotonobetaine proceeds by a stereospecific syn pathway.In contrast, all other hydration-dehydrationreactionsexamined thus
far show that, when the proton added or abstracted isbonded to a carbon atom that is adjacent to acarboxylate group, the absolute stereochemistry o
f the resultingtrans
formation is
anti. Crystallographic details
for[(CH
3)
3N-CH
2-CH(OH)-CHD-COOH]
+[AuCl
4]
-are as
follows: space group
P2
12
12
1 (orthorhombic),
a = 10.855(2),
b = 11.678(3),
c = 22.776(6)Å;
Z = 8;
final agreement
factor
for the neutron analysisat 15 K:
R(
Fo) =0.097based on 1140 re
flections with
I >3
fchars/sigma.gi
f" BORDER=0 >(
I).