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Orotidine 5’‐Monophosphate Decarboxylase: Effect of an S154A Mutation on Enzyme‐Catalyzed Decarboxylation of OMP and on Decarboxylation of Complexes that Contain the Substrate Pieces
Author(s) -
Barnett Shonoi A,
Wood Bryant M,
Gerlt John A,
Richard John P
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.611.7
Subject(s) - decarboxylation , chemistry , substrate (aquarium) , stereochemistry , enzyme , catalysis , biochemistry , biology , ecology
Orotidine 5’‐monophosphate decarboxylase (OMPDC) catalyzes the chemically difficult decarboxylation of orotidine 5’‐monophosphate (OMP) to give uridine 5’‐monophosphate (UMP) (Scheme 1). The ratio of the second order rate constants for decarboxylation of OMP and of a truncated substrate 1‐(β‐D‐erythrofuranosyl)orotic acid (EO), (4 × 10 8 )‐fold, gives an intrinsic phosphate binding energy of 11.8 kcal/mol. More than half of the dianion binding energy (7.8 kcal/mol) can be recovered in the binding of phosphite dianion to the (OMPDC•EO) ‡ complex. The effect of the S154A mutation of the yeast enzyme on the stability of the transition state generated from free OMPDC and OMP (5.7 kcal/mol) is similar to the effect of the mutation on the transition state generated from free OMPDC, EO and phosphite (6.4 kcal/mol), so that the whole substrate and substrate in pieces react through similar transition states. The effect S154A mutation on the reaction of the pieces has been partitioned into a 3.2 kcal/mol effect on the reaction of EO alone, and a 3.1 kcal/mol effect on phosphite activation. The mutation reduces phosphite activation, even though there are apparently no direct interactions between the excised ‐CH 2 OH and the phosphate of OMP. We propose that dianion binding drives loop closure around the transition state, which serves to strengthen the network of hydrogen bonding interactions with the ‐CH 2 OH side chain.