Synthesis of FPP Fluorescent Analogues to Probe the Interactions with Undecaprenyl Pyrophosphate Synthase
Date Issued
2010
Date
2010
Author(s)
Chang, Shu-Lan
Abstract
Undecaprenyl pyrophosphate synthase (UPPs) catalyzes the reactions of eight isopentenyl pyrophosphate (IPP) with farnesyl pyrophosphate (FPP) to generate C55 undecaprenyl pyrophosphate (UPP), which is a carrier transporting lipid II across the membrane in the biosynthesis of bacteria cell wall. Four FPP fluorescent analogues are designed to aim for probing the biosynthetic pathway of bacterial cell wall. It is hoped that these FPP analgues can bind with UPPs as its natural substrate, and follow the original biosynthetic pathway. It is also hoped that these FPP analogues have fluorescence resonance energy transfer phenomena to Alexa Fluoro 488 that has been modified in the active site of UPPs when the chain elongation reaction occurs. In addition, we can apply the fluorescence property for probing the position of the biosynthetic product.
These FPP fluorescent analogues, denoted as BD-GPP、DC-GPP、3HF-GPP and GPP-3HF are composed of fluorophores and geranyl pyrophosphate (GPP) (see figure). Three fluorophores which emit at 500 nm are chosen. One of them is boron-dipyrromethene (BODIPY). The framework of BODIPY connected with geranyl pyrophosphate is similar to UPPs natural substrate, and its high quantun yield and photostability motivate us to apply it to our probe design. The second fluorophore is called dicyanomethylenedihydrofuran (DCDHF). The push-pull structural characteristics renders modification of its photophysical properties possible, and its amino group can be derived easily. The third fluorophore is 3-hydroxyflavone, which has dual emissions and the ratios of the two emissions depending on the polarity of microenvironment. This property that one fluorophore has dual emission stimulates us to apply this fluorophore to this system. After testing these four FPP analogues binding and biological properties activity with UPPs, we find these four FPP analogues can not proceed chain elongation reactions when bound in the active site of UPPs, but all of them can be reasonably good inhibitors.
Subjects
FPP fluorescent analogues
Type
thesis
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