Pyrene·Pyrene Complexes at the Active Site of Cytochrome P450 3A4:  Evidence for a Multiple Substrate Binding Site

2002 ◽  
Vol 124 (40) ◽  
pp. 11866-11867 ◽  
Author(s):  
Michael J. Dabrowski ◽  
Michael L. Schrag ◽  
Larry C. Wienkers ◽  
William M. Atkins
Keyword(s):  
Cytochrome P450 ◽  
Binding Site ◽  
Active Site ◽  
Substrate Binding ◽  
Cytochrome P450 3A4 ◽  
Substrate Binding Site ◽  
Multiple Substrate

scholarly journals Multiple substrate-binding sites are retained in cytochrome P450 3A4 mutants with decreased cooperativity

Xenobiotica ◽  
2010 ◽  
Vol 41 (4) ◽  
pp. 281-289 ◽  
Author(s):  
Harshica Fernando ◽  
Jessica A. O. Rumfeldt ◽  
Nadezhda Y. Davydova ◽  
James R. Halpert ◽  
Dmitri R. Davydov
Keyword(s):  
Cytochrome P450 ◽  
Binding Sites ◽  
Substrate Binding ◽  
Cytochrome P450 3A4 ◽  
Multiple Substrate ◽  
Substrate Binding Sites

Crystal structures of substrate-bound chitinase from the psychrophilic bacteriumMoritella marinaand its structure in solution

2014 ◽  
Vol 70 (3) ◽  
pp. 676-684 ◽  
Author(s):  
Piotr H. Malecki ◽  
Constantinos E. Vorgias ◽  
Maxim V. Petoukhov ◽  
Dmitri I. Svergun ◽  
Wojciech Rypniewski
Keyword(s):  
Binding Site ◽  
Crystal Structures ◽  
Domain Structure ◽  
Active Site ◽  
Substrate Binding ◽  
Glycoside Hydrolases ◽  
Substrate Binding Site ◽  
Domain Structures ◽  
Structure In Solution

The four-domain structure of chitinase 60 fromMoritella marina(MmChi60) is outstanding in its complexity. Many glycoside hydrolases, such as chitinases and cellulases, have multi-domain structures, but only a few have been solved. The flexibility of the hinge regions between the domains apparently makes these proteins difficult to crystallize. The analysis of an active-site mutant ofMmChi60 in an unliganded form and in complex with the substrates NAG4and NAG5revealed significant differences in the substrate-binding site compared with the previously determined complexes of most studied chitinases. A SAXS experiment demonstrated that in addition to the elongated state found in the crystal, the protein can adapt other conformations in solution ranging from fully extended to compact.

scholarly journals Mapping the Substrate Binding Site of Phenylacetone Monooxygenase from Thermobifida fusca by Mutational Analysis

2011 ◽  
Vol 77 (16) ◽  
pp. 5730-5738 ◽  
Author(s):  
Hanna M. Dudek ◽  
Gonzalo de Gonzalo ◽  
Daniel E. Torres Pazmiño ◽  
Piotr Stępniak ◽  
Lucjan S. Wyrwicz ◽  
...  
Keyword(s):  
Substrate Specificity ◽  
Binding Site ◽  
Active Site ◽  
Structural Model ◽  
Mutational Analysis ◽  
Substrate Binding ◽  
Thermobifida Fusca ◽  
Content Type ◽  
Substrate Binding Site

ABSTRACTBaeyer-Villiger monooxygenases catalyze oxidations that are of interest for biocatalytic applications. Among these enzymes, phenylacetone monooxygenase (PAMO) fromThermobifida fuscais the only protein showing remarkable stability. While related enzymes often present a broad substrate scope, PAMO accepts only a limited number of substrates. Due to the absence of a substrate in the elucidated crystal structure of PAMO, the substrate binding site of this protein has not yet been defined. In this study, a structural model of cyclopentanone monooxygenase, which acts on a broad range of compounds, has been prepared and compared with the structure of PAMO. This revealed 15 amino acid positions in the active site of PAMO that may account for its relatively narrow substrate specificity. We designed and analyzed 30 single and multiple mutants in order to verify the role of these positions. Extensive substrate screening revealed several mutants that displayed increased activity and altered regio- or enantioselectivity in Baeyer-Villiger reactions and sulfoxidations. Further substrate profiling resulted in the identification of mutants with improved catalytic properties toward synthetically attractive compounds. Moreover, the thermostability of the mutants was not compromised in comparison to that of the wild-type enzyme. Our data demonstrate that the positions identified within the active site of PAMO, namely, V54, I67, Q152, and A435, contribute to the substrate specificity of this enzyme. These findings will aid in more dedicated and effective redesign of PAMO and related monooxygenases toward an expanded substrate scope.

Sign in / Sign up

Export Citation Format

Share Document