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Day 9 - Seeing is believing: Developing chemical probes to image penicillin binding proteins

16/3/2019

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​Haneesh Gangotra, a 3rd year PhD student with the Florey Institute, shares a brief overview on the multidisciplinary techniques he’s used for imaging penicillin binding proteins (PBPs).

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​PBPs are essential enzymes used by bacteria to synthesise their cell wall and are the target of β-lactam antibiotics such as penicillin. Over time however, bacteria have formed new PBPs which resist these drugs and given rise to highly resistant strains such as MRSA.
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​Being a chemist you would think I found the first part the easiest, that is synthesising suitable fluorescent probes which could bind to PBPs, but it was far from trivial! β-lactam’s are incredibly strained molecules and as such are difficult to manipulate. They can also be liable to degradation and this limits the type of chemistry you can apply. 
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Structure of a penicillin derivatised chemical probe
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​Luckily I eventually developed a protocol to work from and was able to successfully take these probes from the bench, apply to S. aureus and actually see them binding to PBPs using microscopy!
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From synthesis and purification to application and imaging of PBPs in S. aureus: Development of a BODIPY based β-lactam chemical probe

​I have recently established a probe for using in Stochastic Optical Reconstruction Microscopy (STORM) which is something we’ve wanted for a while! This will allow crystal clear images of PBP interactions and offer an insight into their precise location during cell wall biosynthesis.
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