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FUNCTIONAL ANALYSIS AND MOLECULAR DOCKING IDENTIFY TWO ACTIVE SHORT‐CHAIN PRENYLTRANSFERASES IN THE GREEN PEACH APHID, M yzus persicae
Author(s) -
Zhang Yong-Lei,
Li ZhengXi
Publication year - 2012
Publication title -
archives of insect biochemistry and physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.576
H-Index - 66
eISSN - 1520-6327
pISSN - 0739-4462
DOI - 10.1002/arch.21032
Subject(s) - prenyltransferase , stereochemistry , chemistry , enzyme , biochemistry , active site , biosynthesis , docking (animal) , medicine , nursing
Short‐chain prenyltransferases are responsible for biosynthesis of the C 10 –C 20 precursors of a variety of isoprenoids. We previously isolated two different short‐chain prenyltransferases from the green peach aphid, M yzus persicae ( M p IPPS 1 and M p IPPS 2). In this study, the activity of the two aphid prenyltransferases was analyzed in vitro. Kinetic analysis using recombinant enzymes showed that both prenyltransferases could efficiently catalyze the formation of C 10 geranyl diphosphate ( GPP ) and C 15 farnesyl diphosphate ( FPP ) from the C 5 substrates isopentenyl diphosphate ( IPP ) and dimethylallyl diphosphate ( DMAPP ), and M p IPPS 2 had higher catalytic activity than M p IPPS 1. Product analysis by gas chromatography–mass spectrometry demonstrated that FPP was generated as the major product, but GPP could be detected at low enzyme concentrations. Molecular docking revealed that M p IPPS 2 had higher binding affinity with the substrates DMAPP , IPP , and GPP than M p IPPS 1, which supported the experimentally determined kinetic parameters. Molecular docking also identified an amino acid residue ( K 266) critical to the catalytic activity of both M p IPPS 1 and M p IPPS 2. This prediction was subsequently confirmed by site‐directed mutagenesis, in which a point mutation ( K 266I) abolished the activity of both M p IPPS 1 and M p IPPS 2. Our data illustrate that both aphid short‐chain prenyltransferases are active forms, which is in contrast to the previously reported results.