Methylamine adenine dinucleotide (MAD): a co-factor to turn alcohol dehydrogenases into aldolases

2002 ◽  
Vol 80 (6) ◽  
pp. 665-670 ◽  
Author(s):  
Denis Wahler ◽  
Jean-Louis Reymond
Keyword(s):  
Alcohol Dehydrogenase ◽  
Active Site ◽  
Primary Amine ◽  
Adenosine Monophosphate ◽  
Chemoenzymatic Synthesis ◽  
Penicillin G ◽  
Alcohol Dehydrogenases ◽  
Horse Liver Alcohol Dehydrogenase ◽  
Liver Alcohol Dehydrogenase ◽  
Horse Liver

Methylamine adenine dinucleotide (MAD) was prepared in seven steps and 15.3% overall yield from 1-acetoxy-2,3,5-tri-O-benzoyl-D-ribose by chemoenzymatic synthesis. The key step was the phosphate–phosphate coupling between adenosine monophosphate and 2,5-anhydro-1-deoxy-1-phenylacetamide-6-phosphate-D-allitol (3) mediated by carbonyl diimidazole (CDI), followed by the removal of the phenylacetamido group by penicillin G acylase. The MAD co-factor provided a primary amine functionality that was suitably positioned to promote the enamine aldolization of 2-oxopropionamide with aldehydes within the active site of alcohol dehydrogenases, which should lead to the transformation of these stereoselective enzymes into aldolases. Preliminary investigations did not reveal any activity with horse liver alcohol dehydrogenase or yeast alcohol dehydrogenase.Key words: aldolases, alcohol dehydrogenases, co-factor enginering, nucleosides, acylases.

scholarly journals Enantioselective affinity labelling of horse liver alcohol dehydrogenase. Correlation of inactivation kinetics with the three-dimensional structure of the enzyme

1983 ◽  
Vol 211 (2) ◽  
pp. 391-396 ◽  
Author(s):  
K H Dahl ◽  
H Eklund ◽  
J S McKinley-McKee
Keyword(s):  
Alcohol Dehydrogenase ◽  
Active Site ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Inactivation Kinetics ◽  
Horse Liver Alcohol Dehydrogenase ◽  
Three Dimensional Structure ◽  
Liver Alcohol Dehydrogenase ◽  
Horse Liver ◽  
Affinity Labelling

Kinetic data for the inactivation of horse liver alcohol dehydrogenase with S-2-chloro-3-(imidazol-5-yl)propionate at pH8.2 were correlated with the three-dimensional structure of the enzyme. The R-2-chloro-3-(imidazol-5-yl)propionate enantiomer did not inactivate the enzyme, and the reaction is thus enantioselective. Inactivation follows an affinity-labelling mechanism where a reversible complex is formed before the irreversible alkylation and inactivation of the enzyme. A reversible complex is also formed with the non-inactivating enantiomer, and this shows that the selectivity occurs at the irreversible step. By using a computer-controlled display system, models of the two enantiomers of 2-chloro- and 2-bromo-3-(imidazol-5-yl)propionate were built into a model of the enzyme so that the imidazole moiety was liganded to the active-site metal, while the carboxylate group interacted with the general anion-binding site. The conformation of the imidazole derivatives and their orientation in the active site were adjusted to minimize unfavourable steric interactions. It was clear that alkylation of cysteine-46 could proceed with the S-enantiomer bound in this way, but not with the R-enantiomer. Model building thus agrees with the inactivation kinetics and indicates the structural origin of the enantioselectivity.

Sign in / Sign up

Export Citation Format

Share Document