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Cristina Azevedo on her recent ACS Chemical Biology paper

We've asked Cristina Azevedo from the Saiardi Lab to tell us about her recent publication in ACS Chemical Biology, a collaborative effort between the Saiardi Lab and the Jessen Lab at University of Freiburg, Germany.

What discoveries led you to the research described in your publication?

A few years ago, we discovered a new post-translational modification of proteins, that we named polyphosphorylation. As the name indicates, this modification involves phosphate being attached to proteins, but it differs from the canonical protein phosphorylation in four main aspects:
(1) The donor of phosphate is not ATP but inorganic polyphosphate; a liner molecule that is solely composed of orthophosphate residues (imagine ATP but without the adenosine). PolyP exists in all organisms and can range from three to hundreds of phosphates in length.

(2) The modification does not occur on a serine, tyrosine or threonine but on lysine residues
.
(3) More than two phosphates are transferred to the lysine residue/s, hence the name "polyphosphorylation".
(4) This mechanism occurs non-enzymatically
.
We originally studied polyphosphorylation in yeast and identified two target proteins: Nsr1 (a protein involved in ribosomal RNA biogenesis) and Topoisomerase (an enzyme that unwinds super coiled DNA).
 

What were you trying to understand?

After having identified Nsr1 and Top1 we were very interested to understand how wide spread this mechanism was, how many proteins were phosphorylated and whether this mechanism was conserved in other species? Moreover, we wanted to develop tools that would allow us and others to study polyphosphorylation in a more systematic and easy way. So, we collaborated with an organic chemistry lab to develop a polyP molecule that we could easily visualize. We created a molecule with a defined number of phosphates, 8 phosphates, and that is capped at both ends with biotin. Using this chemical, we screened a human proteome library to look for targets of polyphosphorylation.

 

Why is this important?

It is very important because after the genome era we came to realize that the human organism is a lot more complex than once thought. The levels of regulation at protein level are ever more complex with many new post-translation modifications being discovered. So discovering a new one and trying to understand its function is very exciting and important.

 

Can you use an analogy to help me understand your work?
I often talk about cells as being factories (which they obviously are), specially like Japanese factories where everything and everyone has a well-established function in a particular place within the factory and well sign-posted. In a Toyota factory, the person in charge to put the tyres on a car, works in a particular space, wears a particular suit with a colour that defines his/her function. The same happens with proteins. Proteins are the factory workers, and for these and other proteins in the cell to know their function, or where in the cell they need to be or even with whom they should associate with, they need to be identified as such. And this is where post-translation modifications come. They "label" the proteins assigning them certain functions, places in the cell, or associated partners. To understand how the factory works we need to know all the colours and all the suits that workers have.

 
What questions remain to be asked?

So so many. We still know very little about the how this mechanism is regulated. Even though we did a screen in mammalian cells, we believe that the proteins we identified are just a snap-shot of the real number of targets. We also do not know the enzymology of synthesis of polyP in mammalian cells and so it will be fundamental to understand how polyP is produced and localized in higher organisms.

 

Link to Cristina's publication here.

Written by Cristina Azevedo