Protein science conference

During his time at APAF, Herbert had the good fortune to meet and collaborate with a true pioneer of proteomics technology, Professor Pier Giorgio Righetti, then at the University of Verona. Righetti is also speaking at the Asia Oceania Human Proteome Organisation (AOHUPO)/Pacific Rim International Conference on Protein Science in Cairns in June.

In the world of proteomics, fractionation of complex samples has been one of the biggest technological breakthroughs of the last 15 years. At the time, Herbert thought fractionation by isoelectric point looked like an excellent way to break up those samples and run the fractions on narrow pH gradients.

"I had always been a huge fan of Righetti's work because he was so prolific and had been an inventor of so many of the technologies that were out there," Herbert says. "It turned out one of those was a liquid phase isoelectric focusing system called a multi-compartment electrolyser. I read about these things and decided that what it had been built to do was purify individual proteins to complete homogeneity for NMR or crystallography, but it seemed to me that you could equally use them to fractionate complex mixtures."

Herbert organised for Righetti to come out to Macquarie to speak at a meeting he and Brad Walsh were organising. From then on, Herbert and Righetti have worked in close collaboration and have published widely together, predominantly on 2D electrophoresis and isoelectric focusing.

At AOHUPO, Herbert will discuss, of all things, fungal infections. Herbert and his new team at UTS are working with Associate Professor Dee Carter from the University of Sydney, assisting them understand the proteomics of fungal infection, using Cryptococcus gattii as a model. C. gattii is not a common infection in humans, although there has been an outbreak in Vancouver recently, but it is most definitely a problem for koalas, as it lives in eucalyptus trees.

"Our real involvement in this is working with how to break open Crypto and get protein out of it. It's got an incredibly and extremely sticky capsule around the cell which is involved in the virulence of the organism. The capsule actually gets bigger when it is in a lung because that is a more stressful situation for the fungus to be in.

"It's been one of the most difficult organisms we've ever worked on, trying to figure out how to break it open and how to break the sticky interaction between the proteins and this capsule. What happens is if you are not very careful all of the proteins are lost and you actually just spin them out in the centrifuge and chuck them away, thinking it is cell debris."

Herbert's team has hit on an old-fashioned but seemingly forgotten method of breaking these interactions and preparing pure samples - salt. "I wanted to test which salts work the best. We've looked at monovalent and divalent cations and found that lithium chloride works better than anything else. I think that is because it is the smallest atom and therefore you have the highest charge density and you can get it into small areas. And we find it is a really good stripping agent, it gets in and dissociates protein from the capsule and we can then recover significant amounts of protein from Cryptococcus."