MANDELATE RACEMASE MUTANT D270N CO-CRYSTALLIZED WITH (S)-ATROLACTATE

Mechanism of the reaction catalyzed by mandelate racemase: structure and mechanistic properties of the D270N mutant.

Mechanism of the Reaction Catalyzed by Mandelate Racemase. 2. Crystal Structure of Mandelate Racemase at 2.5-A Resolution: Identification of the Active Site and Possible Catalytic Residues

Mandelate Racemase and Muconate Lactonizing Enzyme are Mechanistically Distinct and Structurally Homologous

MSEVLITGLRTRAVNVPLAYPVHTAVGTVGTAPLVLIDLATSAGVVGHSYLFAYTPVALKSLKQLLDDMAAMIVNEPLAP
VSLEAMLAKRFCLAGYTGLIRMAAAGIDMAAWDALGKVHETPLVKLLGANARPVQAYDSHSLDGVKLATERAVTAAELGF
RAVKTKIGYPALDQDLAVVRSIRQAVGDDFGIMVDYNQSLDVPAAIKRSQALQQEGVTWIEEPTLQHDYEGHQRIQSKLN
VPVQMGENWLGPEEMFKALSIGACRLAMPNAMKIGGVTGWIRASALAQQFGIPMSSHLFQEISAHLLAATPTAHWLERLD
LAGSVIEPTLTFEGGNAVIPDLPGVGIIWREKEIGKYLV

THE INHIBITOR CONSTANT (K SUB I) FOR (S)-ATROLACTATE IS APPROXIMATELY 0.2 MILLIMOLAR; CRYSTALS WERE OBTAINED FROM A SOLUTION THAT CONTAINED 5 MILLIMOLAR INHIBITOR.

THIS IS THE STRUCTURE OF THE MANDELATE RACEMASE MUTANT ASPARTATE 270 -> ASPARAGINE CO-CRYSTALLIZED WITH THE COMPETITIVE INHIBITOR (S)-ATROLACTIC ACID.

RESIDUES LEU 18 - THR 31:
THE ELECTRON DENSITY FOR THE RESIDUES IN THE "FLAP"
REGION CONTAINED MULTIPLE BREAKS. IT IS LIKELY THAT THE
"FLAP" RESIDUES WERE ACTUALLY PRESENT IN MULTIPLE
CONFORMATIONS. HOWEVER, THEY WERE REFINED IN THESE
CONFORMATIONS WITH THEIR OCCUPANCIES SET TO 1.0.
CONSEQUENTLY, THEIR B-FACTORS ARE GREATER THAN THE REST
OF THE PROTEIN. FURTHERMORE, THE LACK OF "ENCLOSING"
ELECTRON DENSITY CAUSED MORE SIGNIFICANT GEOMETRIC
DEVIATIONS DURING REFINEMENT.

ISOMERASE, MANDELATE PATHWAY, ATROLACTATE, MAGNESIUM RACEMASE, RACEMASE

MSEVLITGLRTRAVNVPLAYPVHTAVGTVGTAPLVLIDLATSAGVVGHSYLFAYTPVALKSLKQLLDDMAAMIVNEPLAP
VSLEAMLAKRFCLAGYTGLIRMAAAGIDMAAWDALGKVHETPLVKLLGANARPVQAYDSHSLDGVKLATERAVTAAELGF
RAVKTKIGYPALDQDLAVVRSIRQAVGDDFGIMVDYNQSLDVPAAIKRSQALQQEGVTWIEEPTLQHDYEGHQRIQSKLN
VPVQMGENWLGPEEMFKALSIGACRLAMPDAMKIGGVTGWIRASALAQQFGIPMSSHLFQEISAHLLAATPTAHWLERLD
LAGSVIEPTLTFEGGNAVIPDLPGVGIIWREKEIGKYLV

SITE MTL CONSTITUTES THE DIRECT METAL ION LIGANDS.

SITE ACT CONSTITUTES THE ACID/BASE CATALYSTS RESPONSIBLE FOR PROTON ABSTRACTION AND REDELIVERY IN THE RACEMIZATION REACTION.

SITE CAR CONSTITUTES THE BINDING SITE FOR THE CARBOXYL GROUP OF SUBSTRATE. THESE RESIDUES FUNCTION, ALONG WITH THE METAL ION, AS ELECTROPHILIC CATALYSTS.