Structural Determinants of Enzyme Binding Affinity: The E1 Component of Pyruvate Dehydrogenase from Escherichia coli in Complex with the Inhibitor Thiamin Thiazolone Diphosphate
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- 作者：Palaniappa Arjunan ; Krishnamoorthy Chandrasekhar ; Martin Sax ; Andrew Brunskill ; Natalia Nemeria ; Frank Jordan ; and William Furey
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：March 9, 2004
- 年：2004
- 卷：43
- 期：9
- 页码：2405 - 2411
- 全文大小：265K
- 年卷期：v.43,no.9(March 9, 2004)
- ISSN：1520-4995
文摘
Thiamin thiazolone diphosphate (ThTDP), a potent inhibitor of the E1 component from theEscherichia coli pyruvate dehydrogenase multienzyme complex (PDHc), binds to the enzyme with greateraffinity than does the cofactor thiamin diphosphate (ThDP). To identify what determines this difference,the crystal structure of the apo PDHc E1 component complex with ThTDP and Mg2+ has been determinedat 2.1 Å and compared to the known structure of the native holoenzyme, PDHc E1-ThDP-Mg2+ complex.When ThTDP replaces ThDP, reorganization occurs in the protein structure in the vicinity of the activesite involving positional and conformational changes in some amino acid residues, a change in the Vcoenzyme conformation, addition of new hydration sites, and elimination of others. These changes culminatein an increase in the number of hydrogen bonds to the protein, explaining the greater affinity of theapoenzyme for ThTDP. The observed hydrogen bonding pattern is not an invariant feature of ThDP-dependent enzymes but rather specific to this enzyme since the extra hydrogen bonds are made withnonconserved residues. Accordingly, these sequence-related hydrogen bonding differences likewise explainthe wide variation in the affinities of different thiamin-dependent enzymes for ThTDP and ThDP. Thesequence of each enzyme determines its ability to form hydrogen bonds to the inhibitor or cofactor.Mechanistic roles are suggested for the aforementioned reorganization and its reversal in PDHc E1catalysis: to promote substrate binding and product release. This study also provides additional insightinto the role of water in enzyme inhibition and catalysis.