Molecular mechanisms that regulate polarity and spindle orientation in animals - admin supplement to purchase mass spectrometer

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Project abstract! Funding is requested for the purchase of a Bruker Microflex SMART LS MALDI-TOF mass spectrometer for verifying the reagents we produce. The instrument will be used by my research group and by my colleague Brad Nolen who has submitted a cross-referenced application. In R35GM127092, the parent grant for this supplement application, my laboratory proposes to uncover the mechanism of animal cell polarization using the neuroblast as a model system. A key aim is to use reconstituted purified Par complex proteins to understand the mechanism by which polarity is regulated. While we routinely use gel electrophoresis as a method for determining the identity and purity of our reagents, several features of our system make mass spectrometry an important complement to this standard technique. First, a component of the Par complex, atypical Protein Kinase C, migrates abnormally by gel electrophoresis because of essential post translational modifications. Second, our work requires making various modified forms of Par complex proteins (e.g. truncations) and this instrument will allow us to verify them with high resolution mass information. We expect that the additional information provided by this instrument will have a significant and sustained positive impact on our research program by increasing the reliability and reproducibility of key reagents.