Modulation of the rate, enantioselectivity, and substrate specificity of semisynthetic transaminases based on lipid binding proteins using site directed mutagenesis

Hao Kuang, Ronald R. Davies, Mark D. Distefano

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21 Scopus citations

Abstract

Fatty acid binding proteins are a class of small 15 kDa proteins with a simple architecture that forms a large solvent sequestered cavity. In previous work, we demonstrated that reductive amination reactions could be performed in this cavity by covalent attachment of a pyridoxamine cofactor to the protein. Here, we report the results of experiments in which the position of pyridoxamine attachment has been varied by site directed mutagenesis. The conjugate IFABP-PX60 reacts at least 9.4-fold more rapidly than our original conjugate ALBP-PX, while IFABP-PX72 inverts the enantioselectivity of reactions (compared to ALBP-PX) and IFABP-PX104 displays very selective substrate specificity. These results indicate that site-directed mutagenesis can be used to tune the rate, enantioselectivity, and substrate specificity of semisynthetic transaminases based on fatty acid binding proteins.

Original languageEnglish (US)
Pages (from-to)2055-2060
Number of pages6
JournalBioorganic and Medicinal Chemistry Letters
Volume7
Issue number15
DOIs
StatePublished - Aug 5 1997

Bibliographical note

Funding Information:
Acknowledgements. We thankD r. C. Friedenfo r providingth ep lasmides ncodingth em utanpt roteinus sed in thiss tudy.M odelingo f IFABP was performebdy M. Silberglitatn dS. Flaim. This work was supportebdy grants from the National ScienceF oundation( NSF-CHE-9506793)a nd the PetroleumR esearchF und (PRF#28140-G4).R . R. Davies was supportedb y a NationalI nstituteso f Health Training Grant (NIH 2T32GM08347-06).

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