Quick Project Snapshot

The mechanism of de novo prion formation

Prion diseases are associated with the conversion of natively-folded prion protein (PrP) into an infectious conformer (a prion), which accumulates in disease and provides a template for the ongoing misfolding and conversion of the normal conformer.  Prions represent a significant health problem for Australia and the world due to limitations on giving blood, use of human derived biological products, sterilisation of surgical instruments and food preparation considerations.

Protein misfolding by cyclic amplification (PMCA) is a cell-free method developed to accelerate replication of the infectious PrP conformer and involves repeated sonication of natively-folded PrP in the presence of an infectious “seed” [1].  Although originally developed as a tool to detect the presence of minute concentrations of prions, it was recently demonstrated that infectious PrP could be detected during PMCA without the need for an infectious seed.  This de novo prion formation was demonstrated using only recombinant PrP expressed from E. coli, RNA and the phospholipid POPG [2].  Understanding of the molecular mechanism underpinning de novo prion generation during PMCA will provide crucial insight into triggers for prion disease.

Sonication (application of ultrasound energy) during PMCA generates free radicals in solution and we can detect these radicals and show they cause numerous post-translational modifications to PrP during PMCA.  We hypothesise that these radicals, combined with the unique physical conditions present during sonication, provide the necessary chemical and structural trigger for PrP oxidation, covalent modification and misfolding into a de novo prion.

This project will characterise the biochemical changes induced by PMCA using a combination of immunodetection (ELISA, western/dot/slot blotting), fluorescence, spectroscopy and mass spectrometry techniques.  The various post-translational modifications to PrP will be correlated with the de novo infectivity of PMCA reaction products using animal and/or cell culture models of prion disease.

 

 

  1. Morales, R.; Duran-Aniotz, C.; Diaz-Espinoza, R.; Camacho, M. V.; Soto, C. Protein misfolding cyclic amplification of infectious prions, Nature Protocols 2012, 7, 1397–1409.
  2. Wang, F.; Wang, X.; Yuan, C-G.; Ma, J. Generating a Prion with Bacterially Expressed Recombinant Prion Protein, Science 2010, 327, 1132–1135.
  3. Makino, K.; Mossoba, M. M.; Riesz, P. Chemical Effects of Ultrasound on Aqueous Solutions.Formation of Hydroxyl Radicals and Hydrogen Atoms, J. Phys. Chem. 198387, 1369–1377.
Profile
HEAD OF LAB
Dr Simon Drew

Biophysics Laboratory

Simon Drew's research aims to characterise the properties of proteins involved in neurodegenerative diseases, including their interaction with metal ions, membranes and free radicals. By first studying model systems in detail, hypotheses are generated regarding biological function that are tested in vitro and in vivo.

His current research themes include:

  1. Functional roles of N-truncated β-amyloid in copper homeostasis
  2. The role of free radicals in creation of infectious prions
  3. Metal bridging interactions in protein aggregation
  4. In vivo near infrared brain imaging

His lab employs a variety of fundamental techniques in physics and chemistry that are complemented with biological models of disease.

All Projects by this Lab

The mechanism of de novo prion formation
CO-HEAD OF DIVISION

Professor Philip Beart

Profile
CO-HEAD OF DIVISION

Prof Colin Masters

Profile

Biophysics Laboratory

1 Projects
Profile
HEAD OF LAB
Dr Simon Drew

Oxidation Biology Unit

0 Projects
Profile
HEAD OF LAB
Prof Ashley Bush

Prana Laboratory

0 Projects
Profile
HEAD OF LAB
Dr Robert Cherny

Neurodegeneration

Scientists in the Neurodegeneration division interrogate how neurones live, die and can be rescued to improve brain function in degenerative conditions such as Parkinson’s and Motor Neuron Diseases. There is no effective treatment for Motor Neurone Disease and the incidence of Parkinson’s Disease is rising alarmingly in our aging community. Gene abnormalities, energy deprivation, toxic rubbish accumulation and inflammation all contribute to a toxic environment for brain cells. Our teams study these events in animal models and cultured cells, with a view to translating knowledge into new therapies for human patients.

All Labs that operate in this Division

Atomic Pathology LaboratoryBiophysics LaboratoryCellular Neurodegeneration LaboratoryCreutzfeldt Jakob Disease Clinical Research GroupMolecular Gerontology LaboratoryMotor Neurone Disease LaboratoryNational Dementia Diagnostics LaboratoryNeurochemistry of Metal IonsNeurogenesis and Neural Transplantation LaboratoryNeuropathology and Neurodegeneration LaboratoryNeuroproteomics and Metalloproteomics LaboratoryNeurotherapeutics LaboratoryOxidation Biology UnitParkinson's Disease LaboratoryPrana LaboratoryPre-clinical Parkinson’s Disease Research LaboratoryPresynaptic Physiology Stem Cells and Neural Development LaboratorySteroid Neurobiology LaboratorySynaptic Neurobiology LaboratoryThe Australian Imaging Biomarker and Lifestyle Study (AIBL)