Structural Rationale for Low-Nanomolar Binding of Transition State Mimics to a Family GH3 -D-G
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- 作者：Maria Hrmova ; Victor A. Streltsov ; Brian J. Smith ; Andrea Vasella ; Joseph N. Varghese ; and Geoffrey B. Fincher
- 刊名：Biochemistry
- 出版年：2005
- 出版时间：December 20, 2005
- 年：2005
- 卷：44
- 期：50
- 页码：16529 - 16539
- 全文大小：538K
- 年卷期：v.44,no.50(December 20, 2005)
- ISSN：1520-4995
文摘
The interactions of a transition state mimic anilinomethyl glucoimidazole (AmGlcIm), with aKi constant of 0.6 × 10-9 M and a Gibbs free energy value of -53.5 kJ/mol, with a family GH3 
mages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-D-glucan glucohydrolase from barley have been analyzed crystallographically and by ab initio quantummechanical modeling. AmGlcIm binds 3 times more tightly to the mages/gifchars/beta2.gif" BORDER=0 ALIGN="middle">-D-glucan glucohydrolase than apreviously investigated phenyl glucoimidazole. In the enzyme-AmGlcIm complex, an additional residue,Tyr253, and a water molecule positioned between subsites -1 and +1 are recruited for binding. Analysesof the two binary complexes reveal the following. (i) An intricate network exists in which hydrogenbonds between the enzyme's catalytic pocket residues Lys206, His207, Tyr253, Asp285, and Glu491 andthe glucoimidazoles are shorter by 0.15-0.53 Å, compared with distances of hydrogen bonds in theMichaelis complex. (ii) The "glucose" moiety of the glucoimidazoles adopts a 4E conformation that isvital for the low-nanomolar binding. (iii) The N1 atoms of the glucoimidazoles are positioned nearlyoptimally for in-line protonation by the Omages/gifchars/epsilon.gif" BORDER=0 >1 atom of the catalytic acid/base Glu491. (iv) The enzymederives binding energies from both glycone and aglycone components of the glucoimidazoles. (iv) Theprevalent libration motion of the two domains of the enzyme could play a significant role during inducedfit closure in the active site. (v) Modeling based on the structural data predicts that protons could bepositioned on the N1 atoms of the glucoimidazoles, and the catalytic acid/base Glu491 could carry anoverall negative charge. (vi) The enzyme-AmGlcIm complex reveals the likely structure of an earlytransition state during hydrolysis. Finally, the high-resolution structures enabled us to define minimalstructures of oligosaccharides attached to Asn221, Asn498, and Asn600 N-glycosylation sites.