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Mechanism of substrate dephosphorylation in low M r protein tyrosine phosphatase
Author(s) -
Kolmodin Karin,
Nordlund Pär,
Åqvist Johan
Publication year - 1999
Publication title -
proteins: structure, function, and bioinformatics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.699
H-Index - 191
eISSN - 1097-0134
pISSN - 0887-3585
DOI - 10.1002/(sici)1097-0134(19990815)36:3<370::aid-prot11>3.0.co;2-9
Subject(s) - dephosphorylation , chemistry , protonation , reaction mechanism , substrate (aquarium) , protein tyrosine phosphatase , enzyme , valence (chemistry) , transition state , stereochemistry , phosphatase , computational chemistry , ion , biochemistry , organic chemistry , catalysis , oceanography , geology
Substrate dephosphorylation by the low molecular weight protein tyrosine phosphatases proceeds via nucleophilic substitution at the phosphorous atom yielding a cysteinyl phosphate intermediate. However, several questions regarding the exact reaction mechanism remain unanswered. Starting from the crystal structure of the enzyme we study the energetics of this reaction, using the empirical valence bond method in combination with molecular dynamics and free energy perturbation simulations. The free energy profiles of two mechanisms corresponding to different protonation states of the reacting groups are examined along stepwise and concerted pathways. The activation barriers calculated relative to the enzyme‐substrate complex are very similar for both monoanionic and dianionic substrates, but taking the substrate binding step into account shows that hydrolysis of monoanionic substrates is strongly favored by the enzyme, because a dianionic substrate will not bind when the reacting cysteine is ionized. The calculated activation barrier for dephosphorylation of monoanionic phenyl phosphate according to this novel mechanism is 14 kcal mol −1 , which is in good agreement with experimental data. Proteins 1999;36:370–379. © 1999 Wiley‐Liss, Inc.