Nanopore analysis of protein structure and folding

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In a healthy cell, most proteins fold into a specific structure or conformation soon after they are synthesized. When no longer needed, the proteins break down and the amino acid building blocks are reused to make new proteins. Under certain pathological conditions, the normal process of synthesis and renewal malfunctions and misfolded proteins can begin to accumulate. Misfolded proteins are unusual because they are difficult to degrade and their presence eventually interferes with critical cell functions and the cell dies. The misfolded proteins often aggregate into "amyloid plaques", which can be identified under a microscope. The presence of amyloid plaques in the brain is the hallmark of many neurodegenerative diseases, such as Alzheimer's and Parkinson's. Although the proteins that cause these diseases are known, the reasons why they misfold are not well understood. My NSERC research program uses nanopore analysis to study the folding and misfolding of proteins. In this technique, a small pore of nanometre dimensions is inserted into a membrane. A current will flow when a voltage is applied across the pore; if a small protein enters the pore, the current will be blocked for as long as a microsecond and this event can be detected electronically. We have found that the signal caused by the protein is very sensitive to its precise conformation as it enters the pore. For example, we have discovered that a-synuclein, which is involved in Parkinson's disease, folds into a stable conformation upon binding nicotine. This may explain why cigarette smokers have a reduced risk for developing the disease. Over the next 5 years, my short-term objectives focus on the use of three different research approaches (development of a new type of pore, use of alternating current fields to study protein conformation, and analysis of model proteins) to gather detailed structural information. My long-term goal is to understand the factors that influence these conformational changes, as this knowledge may lead to the development of drugs that could be used to treat neurodegenerative and other protein misfolding diseases.