The Mitochondrial Protein Partnership (MPP) is one of the twelve
PSI:Biology High-Throughput Enabled Structural Biology Partnerships funded by the National Institute of General Medical Sciences.
Under the leadership of John Markley, PhD (left), Steenbock Professor of Biomolecular Structure, and David Pagliarini, PhD (right), Assistant Professor, both at the University of Wisconsin-Madison, the MPP is dedicated to studying the role of mitochondria in human health and disease.
To tackle this project, Professors Markley and Pagliarini have assembled a multidisciplinary team of investigators that includes structural biologists, enzymologists, and
mitochondrial researchers. The experience of this team of researchers at the University of Wisconsin-Madison, many of whom were involved in the PSI-funded Center for Eukaryotic Structural Genomics (2002-2010), has uniquely positioned the MPP to apply PSI-developed technologies to chosen
mitochondrial targets. “We became interested in the mammalian mitochondrial proteome because there are so many fundamental questions yet to be answered, and because of
their profound role in disease pathophysiology. Among our top 1200 proteins of highest interest, hundreds have not been
characterized using biological methods,” said Markley. “Furthermore, mitochondrial dysfunction has been implicated in more
than 50 human diseases, such as type II diabetes and cancer.”
MPP’s target list includes the full mitochondrial proteomes from human, mouse, and other model eukaryotes, but priority is given to targets nominated by members of the mitochondrial research community that meet the goals of the MPP. Accepted targets are provided to the Northeast Structural Genomics Consortium (NESG), the High-throughput PSI:Biology Center partnered with the MPP, which puts them into their structure determination pipeline. At the MPP, the accepted targets are screened by an automated in vitro protein production system that utilizes a cell-free wheat germ extract developed by CellFree Sciences with CESG’s help. Since its development, the cell-free system has been used to produce labeled samples (both NMR and x-ray crystallography) and membrane protein complexes on a large scale. In addition to the cell-free system, the MPP uses an E. coli cell-based platform developed by the CESG. The MPP uses these approaches to produce proteins for biological, biochemical, and biophysical studies.
The relationship with NESG benefits both the MPP investigators and their collaborators. The MPP works with the NESG to determine which approach (cell-based or cell-free) is best for producing protein for structural and functional investigations. To date, six structures have been deposited into the PDB by the NESG and MPP, including the mitochondrial succinate dehydrogenase assembly factor 2 from S. cerevisiae, shown here. Markley stated, “In conjunction with the NESG and our collaborators, we are solving structures and designing appropriate functional follow-up studies so that we can construct complete structure-function stories for each target.”
To learn more about these proteins and their role in disease, the Partnership applies multiple techniques to study protein-protein and protein-small molecule interactions, protein dynamics and conformational transitions, as well as enzymatic and mitochondrial import activity. “When we provide a key link between a structure and its biomedical impact,” said Markley, “we are accomplishing our main goal - to elucidate the role of mitochondria in human health and disease.”
Mitochondrial Protein Partnership researchers at University of Wisconsin-Madison.
Posted September 2012.